ascorbic-acid and epigallocatechin-gallate

It is thought that germ cells and preimplantation embryos during development are most susceptible to endogenous and exogenous environmental factors because the epigenome in those cells is undergoing dramatic elimination and reconstruction. Exposure to environmental factors such as nutrition, climate, stress, pathogens, toxins, and even social behavior during gametogenesis and early embryogenesis has been shown to influence disease susceptibility in the offspring. Early-life epigenetic modifications, which determine the expression of genetic information stored in the genome, are viewed as one of the general mechanisms linking prenatal exposure and phenotypic changes later in life. From atmospheric pollution, endocrine-disrupting chemicals to heavy metals, research increasingly suggests that environmental pollutions have already produced significant consequences on human health. Moreover, mounting evidence now links such pollution to relevant modification in the epigenome. The epigenetics diet, referring to a class of bioactive dietary compounds such as isothiocyanates in broccoli, genistein in soybean, resveratrol in grape, epigallocatechin-3-gallate in green tea, and ascorbic acid in fruits, has been shown to modify the epigenome leading to beneficial health outcomes. This review will primarily focus on the causes and consequences of prenatal environment pollution exposure on the epigenome, and the potential protective role of the epigenetics diet, which could play a central role in neutralizing epigenomic aberrations against environmental pollutions.

Topics: Ascorbic Acid; Brassica; Catechin; Diet; Environmental Pollutants; Epigenesis, Genetic; Female; Genistein; Humans; Plant Extracts; Pregnancy; Protective Agents; Resveratrol

Polyphenols, found abundantly in plants, display many anticarcinogenic properties including their inhibitory effects on cancer cell proliferation, tumor growth, angiogenesis, metastasis, and inflammation as well as inducing apoptosis. In addition, they can modulate immune system response and protect normal cells against free radicals damage. Most investigations on anticancer mechanisms of polyphenols were conducted with individual compounds. However, several studies, including ours, have indicated that anti-cancer efficacy and scope of action can be further enhanced by combining them synergistically with chemically similar or different compounds. While most studies investigated the anti-cancer effects of combinations of two or three compounds, we used more comprehensive mixtures of specific polyphenols and mixtures of polyphenols with vitamins, amino acids and other micronutrients. The mixture containing quercetin, curcumin, green tea, cruciferex, and resveratrol (PB) demonstrated significant inhibition of the growth of Fanconi anemia head and neck squamous cell carcinoma and dose-dependent inhibition of cell proliferation, matrix metalloproteinase (MMP)-2 and -9 secretion, cell migration and invasion through Matrigel. PB was found effective in inhibition of fibrosarcoma HT-1080 and melanoma A2058 cell proliferation, MMP-2 and -9 expression, invasion through Matrigel and inducing apoptosis, important parameters for cancer prevention. A combination of polyphenols (quercetin and green tea extract) with vitamin C, amino acids and other micronutrients (EPQ) demonstrated significant suppression of ovarian cancer ES-2 xenograft tumor growth and suppression of ovarian tumor growth and lung metastasis from IP injection of ovarian cancer A-2780 cells. The EPQ mixture without quercetin (NM) also has shown potent anticancer activity in vivo and in vitro in a few dozen cancer cell lines by inhibiting tumor growth and metastasis, MMP-2 and -9 secretion, invasion, angiogenesis, and cell growth as well as induction of apoptosis. The presence of vitamin C, amino acids and other micronutrients could enhance inhibitory effect of epigallocatechin gallate (EGCG) on secretion of MMPs. In addition, enrichment of NM with quercetin (EPQ mix) enhanced anticancer activity of NM in vivo. In conclusion, polyphenols, especially in combination with other polyphenols or micronutrients, have been shown to be effective against multiple targets in cancer development and progression, and s

Topics: Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Ascorbic Acid; Biological Availability; Catechin; Cell Line, Tumor; Cell Proliferation; Curcumin; Disease Models, Animal; Humans; Micronutrients; Neoplasms; Plant Extracts; Polyphenols; Quercetin; Resveratrol; Stilbenes; Tea

Flavonoids are described to exert a large array of biological activities, which are mostly ascribed to their radical-scavenging, metal chelating and enzyme modulation ability. Most of these evidences have been obtained by in vitro studies on individual compounds and at doses largely exceeding those dietary. Little is known about a possible relationship between rate and extent of the absorption and modifications of plasma antioxidants. To elucidate this aspect, human volunteers were supplemented with single doses of green tea catechins in free (Greenselect) or phospholipid complex form (Greenselect Phytosome) equivalent to 400 mg epigallocatechingallate (EGCg). EGCg was chosen as biomarker for green tea catechin absorption, and its time course plasma concentration was correlated to the subsequent percent variations of plasma ascorbate, total glutathione, alpha-tocopherol, beta-carotene and Total Radical Antioxidant Parameter (TRAP). Green tea catechins were absorbed more extensively when administered as phospholipid complex rather than as free catechins. Single dose intake of both forms of catechins produced a transient decrease (10-20%) of plasma ascorbate and total glutathione and an increase of plasma TRAP (16-19%). These variations were consistent with the plasmatic levels of EGCg, ascorbate and total glutathione.

Topics: Adult; Antioxidants; Ascorbic Acid; beta Carotene; Biomarkers; Catechin; Dietary Supplements; Glutathione; Humans; Male; Phospholipids; Tea; Time Factors; Vitamin E

Methamphetamine (METH) use disorder is a chronic, relapsing disorder and involves frequent failures of self-control of drug seeking and taking. Epigallocatechin-3-gallate (EGCG) is the most abundant polyphenolic compounds of green tea, which has shown great therapeutic effectiveness in neurological disorders. However, it is still unknown whether and how EGCG affects METH seeking behaviour. Here, we show nanostructured EGCG/ascorbic acid nanoparticles (EGCG/AA NPs) dose-dependently reduced METH self-administration (SA) under fixed-ratio 1 (FR1) and progressive ratio (PR) reinforcement schedules in mice and shifted METH dose-response curves downward. Furthermore, EGCG/AA NPs decreased drug- and cue-induced METH seeking. In addition, we found that METH SA led to a decrease in inhibitory postsynaptic currents (IPSCs) and increase in the AMPAR/NMDAR ratio and excitation/inhibition (E/I) ratio in ex vivo midbrain slices from ventral tegmental area (VTA) dopamine neurons. EGCG/AA NPs enhanced Gamma-aminobutyric acid (GABA)ergic inhibition and normalized the E/I ratio. EGCG restored the balance between excitation and inhibition in VTA dopamine neurons, which may contribute to the attenuation of METH SA. These findings indicate that EGCG is a promising pharmacotherapy for METH use disorder.

Topics: Animals; Ascorbic Acid; Catechin; Drug-Seeking Behavior; Methamphetamine; Mice; Reinforcement Schedule; Self Administration

A series of incubation systems of pure (-)-Epigallocatechin gallate (EGCG), ascorbic acid (AA) and dehydroascorbic acid (DHAA) at 80 °C were performed to investigated the effect and mechanism of AA on the stability of EGCG. Results shows the dual function of AA, protect action at low concentration and promoting degradation at high concentration, and the critical concentration is about 10 mmol/L. The protective properties of AA due to the reversible reaction from AA to DHAA inhibiting oxidation pathway of EGCG to EGCG quinone or other activated intermediates, and both AA and DHAA can inhibit the hydrolysis of EGCG. The properties of promoting degradation is mainly due to the fact that DHAA, the oxidation product of AA, can react with EGCG to generate some ascorbyl adducts of EGCG. This result is helpful to control the stability of catechins and further clarify the complex interaction on healthy between EGCG and AA.

Topics: Ascorbic Acid; Catechin; Dehydroascorbic Acid; Hydrolysis; Oxidation-Reduction; Tea; Temperature

Alzheimer's disease (AD) is a progressive neurodegenerative disease that is exacerbated by social isolation (SI) and protein malnutrition (PM). Antioxidants, physical and mental activities (Ph&M) can maintain cognitive functions and protect against dementia.. To investigate the impact of Epigallocatechin-3-gallate (EGCG), Vitamin E (VE), Vitamin C (VC), and Selenium (Se), in enhancing the potential effect of Ph&M versus SI&PM as risk factors in the progression of AD in rats.. Aluminum chloride (70 mg/kg, I.P for 5 weeks) was used to induce AD in rats that either normally fed or socially isolated and protein malnourished (SI&PM). Simultaneously, rats were weekly exposed to Ph&M either alone or in combination with EGCG (10 mg/kg, I.P), VC (400 mg/kg, P.O), VE (100 mg/kg, P.O), and Se (1 mg/kg, P.O).. The combination protocol of EGCG, VE, VC, and Se together with Ph&M significantly increased brain monoamines, superoxide dismutase (SOD), total antioxidant capacity (TAC) and brain-derived neurotrophic factor (BDNF) in AD, SI&PM and SI&PM/AD groups. Additionally, this regimen significantly mitigated brain acetylcholine esterase (ACHE), β-amyloid (Aβ), Tau protein, β-secretase, malondialdehyde (MDA), tumor necrosis factor-alpha (TNF-α), and Interleukin 1β (IL-1β) as well as DNA fragmentation. These biochemical findings were supported by the histopathological examinations of brain tissue.. The combination protocol of antioxidants with Ph&M activities mitigated SI&PM-induced progressive risk of AD.

Topics: Alzheimer Disease; Animals; Antioxidants; Ascorbic Acid; Catechin; Disease Models, Animal; Disease Progression; Male; Mental Health; Physical Conditioning, Animal; Rats; Rats, Sprague-Dawley; Risk Factors; Selenium; Vitamin E

Epigallocatechin gallate (EGCG) is a potential therapeutic agent for treatment of atopic dermatitis (AD) due to its antioxidant and anti-inflammatory activities. However, inherent instability of EGCG greatly limits its bioavailability and clinical efficacy. In this study, we developed a poly-γ-glutamate (γ-PGA)-based microneedle (MN) formulation capable of maintaining EGCG's stability and efficiently delivering EGCG into the skin to ameliorate AD symptoms. The γ-PGA MN can not only protect EGCG from oxidation, but also serve as an immunomodulator to downregulate T helper type 2 (Th2)-type immune responses. Encapsulation of EGCG into the γ-PGA MN and utilization of L-ascorbic acid (AA) as a stabilizer preserved 95% of its structural stability and retained 93% of its initial antioxidant activity after 4 weeks of storage. Once-weekly administration of EGCG/AA-loaded MNs to an Nc/Nga mouse model of AD for 4 weeks significantly ameliorated skin lesions and epidermal hyperplasia by reducing serum IgE (from 12156 ± 1344 to 5555 ± 1362  ng/mL) and histamine levels (from 81 ± 18 to 40 ± 5 pg/mL) and inhibiting IFN-γ (from 0.10 ± 0.01 to 0.01 pg/mg total protein) and Th2-type cytokine production, when compared to the AD (no treatment) group (p < 0.05). Notably, once-weekly MN therapy was at least as effective as the daily topical application of an EGCG + AA solution but markedly reduced the administration frequency and required dose. These results show that EGCG/AA-loaded γ-PGA MNs may be a convenient and promising therapeutic option for AD treatment. STATEMENT OF SIGNIFICANCE: This study describes epigallocatechin gallate (EGCG)/L-ascorbic acid (AA)-loaded poly-γ-glutamate (γ-PGA) microneedles (MN) capable of providing antioxidant, anti-inflammatory, and immunomodulatory effects on inflamed skin for ameliorating atopic dermatitis (AD) symptoms in Nc/Nga mice. After skin insertion, the γ-PGA MN can be quickly dissolved in the skin and remain in the dermis for sustained release of encapsulated active ingredients for 6 days. We demonstrated that once-weekly MN therapy effectively alleviated skin lesions and modulated immune response to relieve Th2-polarized allergic response in mice. Once-weekly MN dosing regimen may provide patients with a more convenient, therapeutically equivalent option to daily topical dosing, and may increase compliance and long-term persistence with AD therapy.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Ascorbic Acid; Catechin; Cytokines; Dermatitis, Atopic; Humans; Immunity; Mice; Polyglutamic Acid; Skin

The wounds of diabetic patients are difficult to heal, which could lead to a limb amputation or even death. The experiment aims to develop a new type of nanoparticles that could accelerate wound healing. Epigallocatechin gallate, ascorbic acid, gelatin and chitosan nanoparticles (EV NPS) were prepared by ion cross-linking method, and their properties were studied. The optimal formula ratio of EV NPS is Vc:EGCG:Gel:CS = 0.2:3:1:1. Transmission electron microscope (TEM) images show that it is a roughly uniform spherical nanoparticle with a diameter of 200 nm. ICR mice were intraperitoneally injected with streptozocin (STZ) to establish diabetic mice. Full-thickness excisional wounds were established on the back of mice. The results showed that EV NPS can promote wound healing in diabetic mice, and the mechanism may be through increasing collagen accumulation, promoting angiogenesis and reducing the infiltration of inflammatory cells. EV NPS may have potential application values for wound healing in diabetic mice.

Topics: Animals; Ascorbic Acid; Catechin; Chitosan; Collagen; Diabetes Mellitus, Experimental; Gelatin; Male; Mice; Mice, Inbred ICR; Nanoparticles; Streptozocin; Wound Healing

Exposure to cadmium (Cd) is a risk factor to health impairments, wherein its cytotoxicity is attributed to induction of oxidative stress. Usage of anti-oxidants, however, can help lessen the damaging effects of Cd. The effect of Cd interaction with low concentration of dietary anti-oxidants, L-ascorbic acid and (-)-epigallocatechin gallate (EGCG), to PC12 cellular mechanisms was examined. The expected toxicity of Cd was observed on PC12 cells but addition of L-ascorbic acid ameliorated this effect. On the other hand, addition of EGCG was able to increase the cytotoxicity of Cd and to decrease the protective effect of L-ascorbic acid against Cd. Increase in LDH activity and decrease in free sulfhydryl levels indicated cell membrane damage and oxidative stress, respectively, in Cd- and EGCG-Cd-treated cells. Downregulation of pro-apoptotic proteins (pro-caspase-9, p53, and ERK1) was observed in cells treated with Cd alone and EGCG-Cd, while upregulation of autophagy-linked proteins (p62 and pBeclin1) was found on L-ascorbic acid-Cd combination treatments. These findings indicate that Cd causes cells to undergo an autophagy-enhanced cell death; low-concentration EGCG and L-ascorbic acid promotes cell survival individually; however, interaction of EGCG with Cd showed enhancement of Cd toxicity and antagonism of L-ascorbic acid efficiency.

Topics: Animals; Apoptosis; Ascorbic Acid; Cadmium; Catechin; Oxidative Stress; PC12 Cells; Rats; Reactive Oxygen Species

The antioxidant and anti-inflammatory properties of Malpighia emarginata D.C (acerola) and Camellia sinensis L. (green tea) have been studied, particularly as an alternative in medicinal approach for different physio pathological conditions. Here we develop an powder blend formulated with both Malpighia emarginata D.C and Camellia sinensis L. which have in the composition higher content of ascorbic acid and epigallatocathechin-3-gallate respectively. Using different conditions for microencapsulation of biocompounds, we performed the powder production through spray-drying process. After, we evaluate the antioxidant and anti-inflammatory properties of blends formulated with Malpighia emarginata D.C and Camellia sinensis L. in an in vitro model of inflammation, using LPS-stimulated RAW-264.7 macrophage cell line. We observed that co-treatment with blends was able to modulate the redox parameters in cells during the in vitro inflammatory response. Moreover, the co-treatment with blends were able to modulate inflammatory response by altering the secretion of cytokines IL-1β, IL-6, IL-10, and TNF-α. Taken together, our results demonstrate for the first time the synergistic effects antioxidant and anti-inflammatory of Malpighia emarginata D.C and Camellia sinensis L. These results warrant further use of the blend powder for use in the products to heath beneficial, principally in terms of prevention of chronic diseases.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Ascorbic Acid; Camellia sinensis; Catechin; Cytokines; Inflammation; Inflammation Mediators; Lipopolysaccharides; Macrophages; Malpighiaceae; Mice; Plant Extracts; RAW 264.7 Cells

To investigate the neuroprotective effects of six natural compounds (caffeine, gallic acid, resveratrol, epigallocatechin gallate [EGCG], L-ascorbic acid and alpha tocopherol [Vitamin E] on heavy metal-induced cell damage in rat PC12 cells.. In this. The metals decreased cell viability but the natural compounds attenuated their effects on apoptosis, necrosis and reactive oxygen species (ROS) levels. Mitochondrial protein changes were involved in the regulation.. Overall, the natural compounds did provide protection against the metal-induced PC12 cell damage. These data suggest that natural compounds may have therapeutic potential against metal-induced neurodegenerative disease.

Topics: alpha-Tocopherol; Animals; Antioxidants; Apoptosis; Ascorbic Acid; Caffeine; Catechin; Cell Survival; China; Gallic Acid; Heavy Metal Poisoning; Metals, Heavy; Neuroprotective Agents; Oxidative Stress; PC12 Cells; Rats; Reactive Oxygen Species; Resveratrol

Human amniotic fluid-derived mesenchymal stem cells (AF-MSCs) may be potentially applied in cell therapy or regenerative medicine as a new alternative source of stem cells. They could be particularly valuable in restoring cardiac tissue after myocardial infarction or other cardiovascular diseases. We investigated the potential of biologically active compounds, namely, angiotensin II, retinoic acid (RA), epigallocatechin-3-gallate (EGCG), vitamin C alone, and the combinations of RA, EGCG, and vitamin C with angiotensin II to induce cardiomyogenic differentiation of AF-MSCs. We observed that the upregulated expression of cardiac gene markers (NKX2-5, MYH6, TNNT2, and DES) and cardiac ion channel genes (sodium, calcium, the potassium) also the increased levels of Connexin 43 and Nkx2.5 proteins. Extracellular flux analysis, applied for the first time on AF-MSCs induced with biologically active compounds, revealed the switch in AF-MSCS energetic phenotype and enhanced utilization of oxidative phosphorylation for energy production. Moreover, we demonstrated changes in epigenetic marks associated with transcriptionally active (H3K4me3, H3K9ac, and H4hyperAc) or repressed (H3K27me3) chromatin. All in all, we demonstrated that explored biomolecules were able to induce alterations in AF-MSCs at the phenotypic, genetic, protein, metabolic, and epigenetic levels, leading to the formation of cardiomyocyte progenitors that may become functional heart cells in vitro or in vivo.

Topics: Adult; Amniotic Fluid; Angiotensin II; Ascorbic Acid; Calcium Channels; Cardiac Myosins; Catechin; Cell Differentiation; Connexin 43; Epigenesis, Genetic; Female; Histones; Homeobox Protein Nkx-2.5; Humans; Mesenchymal Stem Cells; Myocytes, Cardiac; Myosin Heavy Chains; Oxidative Phosphorylation; Potassium Channels; Pregnancy; Pregnancy Trimester, Second; Primary Cell Culture; Signal Transduction; Tretinoin; Troponin T

Some amino acids have strong antioxidant activity in frying oil. This study aimed to obtain further information including antioxidant activity at different concentrations and interactions with rosemary extract, green tea extract, and ascorbic acid. Antioxidant activity of arginine, cysteine, lysine, methionine, and tryptophan was examined by increasing the concentration in soybean oil (SBO) at 180 °C within the concentration range of 0 to 15 mM. These amino acids showed increased activity with increasing concentration without showing prooxidant activity at the given concentration range. Addition of 15 mM methionine did not inhibit the prooxidant activity of α-tocopherol at high concentrations in SBO while it significantly increased the activity at each concentration of α-tocopherol. Methionine showed an additive effect with a commercial rosemary extract while lysine had an antagonistic interaction in SBO at the total concentration of 5.5 mM. Mixtures of green tea extract and methionine did not show better activity than methionine alone in SBO and stripped SBO. (-)-Epigallocatechin gallate, the major active component in green tea, showed a synergistic effect with methionine in stripped SBO but there was no significant interaction effect in SBO. Although ascorbic acid had a synergistic effect with methionine in stripped SBO, it showed a significant antagonistic effect in SBO. Methionine had strong antioxidant activity in six other vegetable oils showing a moderate correlation (R

Topics: alpha-Tocopherol; Amino Acids; Antioxidants; Ascorbic Acid; Catechin; Cooking; Hot Temperature; Plant Extracts; Rosmarinus; Soybean Oil; Tea; Tocopherols

Topics: Alzheimer Disease; Amyloid beta-Peptides; Animals; Antioxidants; Ascorbic Acid; Brain; Catechin; Disease Models, Animal; Drug Carriers; Drug Liberation; Endothelial Cells; Male; Mice, Inbred C57BL; Mice, Transgenic; Nanoparticles; Polyethylene Glycols; Polylactic Acid-Polyglycolic Acid Copolymer; Rats

Noble metal nanoparticles (NPs) have been widely used in many consumer products. Their effects on the antioxidant activity of commercial dietary supplements have not been well evaluated. In this study, we examined the effects of gold (Au NPs), silver (Ag NPs), platinum (Pt NPs), and palladium (Pd NPs) on the hydroxyl radical (·OH) scavenging ability of three dietary supplements vitamin C (L-ascorbic acid, AA), (-)-epigallocatechin gallate (EGCG), and gallic acid (GA). By electron spin resonance (ESR) spin-trapping measurement, the results show that these noble metal NPs can inhibit the hydroxyl radical scavenging ability of these dietary supplements.

Topics: Antioxidants; Ascorbic Acid; Catechin; Dietary Supplements; Electron Spin Resonance Spectroscopy; Free Radical Scavengers; Gallic Acid; Gold; Hydroxyl Radical; Metal Nanoparticles; Palladium; Platinum; Silver

Light chains are abnormally overexpressed from disordered monoclonal B-cells and form amyloid fibrils, which are then deposited on the affected organ, leading to a form of systemic amyloidosis known as AL (Amyloid Light chain) amyloidosis. A green tea catechin, epigallocatechin-3-O-gallate (EGCG), which is thought to inhibit various amyloidoses, is a potent inhibitor of amyloid fibril formation in AL amyloidosis.. An amyloidogenic variable domain in λ6 light chain mutant, Wil was incubated in the presence of EGCG. The incubation products were analyzed by SDS-PAGE and reverse-phase HPLC. The interaction between Wil and EGCG was observed by using NMR and tryptophan fluorescence.. EGCG inhibited the amyloid fibril formation of Wil at pH 7.5 and 42 °C. Under these conditions, most Wil populations were in the unfolded state and several chemical reactions, i.e., oxidation and/or covalent bond oligomerization could be induced by auto-oxidated EGCG. Moreover, we found that EGCG bound to the unfolded state of Wil with higher affinity (Kd = 7 μM).. Inhibition of amyloid fibril formation of Wil was caused by 1) EGCG binding to unfolded state rather than folded state and 2) chemical modifications of Wil by auto oxidation of EGCG.. In the competitive formation of amyloid fibrils and off-pathway oligomers, EGCG produces the latter immediately after it preferentially binds to the unfolded state. It may be general mechanism of EGCG inhibition for amyloidosis.

Topics: Amino Acid Sequence; Amyloid; Ascorbic Acid; Catechin; Chromatography, High Pressure Liquid; Electrophoresis, Polyacrylamide Gel; Immunoglobulin Light Chains; Methionine; Mutation; Oxidation-Reduction; Peptide Mapping; Protein Binding; Protein Folding; Protein Unfolding; Spectrum Analysis; Thermodynamics

There are large differences between flavonoids to protect against apoptosis, a process in which cytochrome c (Cyt c) plays a key role. In this work, we show that 7 of 13 flavonoids studied have a capacity to reduce Cyt c similar or higher than ascorbate, the flavonols quercetin, kaempferol and myricetin, flavanol epigallocatechin-gallate, anthocyanidins cyanidin and malvidin, and the flavone luteolin. In contrast, the kaempferol 3(O)- and 3,4'(O)-methylated forms, the flavanone naringenin, and also apigenin and chrysin, had a negligible reducing capacity. Equilibrium dialysis and quenching of 1,6-diphenyl-1,3,5-hexatriene fluorescence experiments showed that flavonoids did not interfere with Cyt c binding to cardiolipin (CL)/phosphatidylcholine (PC) vesicles. However, the CL-induced loss of Cyt c Soret band intensity was largely attenuated by flavonoids, pointing out a stabilizing action against Cyt c unfolding in the complex. Moreover, flavonoids that behave as Cyt c reductants also inhibited the pro-apoptotic CL-induced peroxidase activity of Cyt c, indicating that modulation of Cyt c signaling are probable mechanisms behind the protective biological activities of flavonoids. © 2016 BioFactors, 43(3):451-468, 2017.

Topics: Animals; Anthocyanins; Ascorbic Acid; Cardiolipins; Catechin; Cytochromes c; Diphenylhexatriene; Flavonoids; Fluorescent Dyes; Horses; Kaempferols; Luteolin; Oxidation-Reduction; Peroxidases; Phosphatidylcholines; Protein Binding; Protein Conformation; Quercetin; Reducing Agents; Spectrometry, Fluorescence; Static Electricity; Unilamellar Liposomes

In this study, simultaneous electrochemical detection of ascorbic acid (AA), dopamine (DA), and uric acid (UA) was performed using a modified graphite paste electrode (MGPE) with epigallocatechin gallate (EGCG) and green tea (GT) powder. It was shown that the anodic peak current increased in comparison with that of the graphite paste electrode (GPE) in the cyclic voltammograms. The optimal pH for simultaneous determination of a quaternary mixture of AA-DA-UA was determined to be pH 2. The anodic peak potentials for a mixture containing AA-DA-UA were well separated from each other. The catalytic peak currents obtained at the surface of the MGPE/EGCG were linearly dependent on the AA, DA, and UA concentrations up to 23, 14, and 14 µM, respectively. The detection limits for AA, DA, and UA were 190, 90, and 70 nM, respectively. The analytical performance of this sensor has been evaluated for simultaneous detection of AA, DA, and UA in real samples. Finally, a modified electrode was prepared using GT and used for simultaneous determination of AA, DA, and UA. Based on the results, MPGE/GT showed two oxidation peaks at 0.43 and 0.6 V for DA and UA, respectively, without any oxidation peak for AA. The calibration curves at the surface of MGPE/GT were linear up to 14 µM with a detection limit of 0.18 and 0.33 µM for DA and UA, respectively. MGPEs provide a promising platform for the future development of sensors for multiplexed electrochemical detection of clinically important analytes.

Topics: Ascorbic Acid; Biosensing Techniques; Catechin; Electrochemical Techniques; Electrochemistry; Electrodes; Graphite; Oxidation-Reduction; Uric Acid

Epigallocatechin gallate (EGCG), the most abundant flavanoid in green tea, is currently being evaluated in the clinic due to its benefits in the treatment of amyloid disorders. Its anti-amyloidogenic effect has been attributed to direct interaction of the intact molecule with misfolded polypeptides. In addition, antioxidant activity is also involved in the anti-amyloidogenic role. The detailed molecular mechanism is still unclear and requires further investigation. In the present study, the kinetics of EGCG oxidation and the anti-amyloidogenic effect of the resultant oxidation substances have been examined. The results indicate that EGCG degrades in a medium at pH 8.0 with a half-life less than 2h. By utilizing lysozyme as an in vitro model, the oxidized EGCG demonstrates a more potent anti-amyloidogenic capacity than the intact molecule, as shown by ThT and ANS fluorescence, TEM determination, and hemolytic assay. The oxidized EGCG also has a stronger disruptive effect on preformed fibrils than the native form. Ascorbic acid eliminates the disruptive role of native EGCG on the fibrils, suggesting that oxidation is a prerequisite in fibril disruption. The results of this work demonstrate that oxidized EGCG plays a more important role than the intact molecule in anti-amyloidogenic activity. These insights into the action of EGCG may provide a novel route to understand the anti-amyloidogenic activity of natural polyphenols.

Topics: Amyloidogenic Proteins; Animals; Antioxidants; Ascorbic Acid; Catechin; Chickens; Half-Life; Humans; Hydrogen-Ion Concentration; Hydrolysis; Kinetics; Muramidase; Oxidation-Reduction; Solutions

The contribution of (-)-epigallocatechin gallate (EGCg) intake to in vivo antioxidant activity is unclear, even with respect to plasma. In this study, we examined how administration of EGCg contributes to plasma antioxidant activity, relative to its concentration, endogenous antioxidants, and assay methods, namely oxygen radical absorbance capacity (ORAC) and ferric reducing/antioxidant power (FRAP). Administration of EGCg (500 mg/kg) to rats increased plasma EGCg (4μmol/L as free form) and ascorbic acid (1.7-fold), as well as ORAC (1.2-fold) and FRAP (3-fold) values. The increase in plasma ascorbic acid following EGCg administration was accompanied by its relocation from the adrenal glands and lymphocytes into plasma, and was related to the increase in FRAP. Plasma deproteinization and assays in plasma model solutions revealed that protein levels significantly contributed to ORAC values, where <3 μmol/L EGCg in the presence of protein exhibited minimal antioxidant activity, as measured by both FRAP and ORAC. As the concentration of plasma ascorbic acid was not influenced by deproteinization, differences in FRAP values with and without deproteinization were estimated to determine the contribution of enhanced ascorbic acid attributable to EGCg administration. These results will help to understand the points that should be considered when evaluating EGCg antioxidant activity in plasma.

Topics: Animals; Antioxidants; Ascorbic Acid; Blood Chemical Analysis; Catechin; Lymphocytes; Male; Oxidation-Reduction; Plasma; Rats; Rats, Wistar

Dietary lipids containing high concentrations of polyunsaturated fatty acids are considered to be beneficial to human health, yet their incorporation within formulated foods is complicated by their susceptibility to oxidation. Lipid oxidation in foods is inhibited through the incorporation of antioxidants, yet the list of antioxidants approved for food use is small, and consumers frequently demand foods without synthetic additives. As a consequence, food processors are now tasked with improving the efficacy of approved, "natural" (i.e., nonsynthetic) antioxidants; a rational strategy for doing so involves localizing the antioxidants at the interface where oxidation usually occurs and regenerating the consumed antioxidants after the oxidation event has occurred. The present study describes a procedure to evaluate antioxidant interactions in oil-in-water food emulsions, which is based on controlled oxidation reactions induced in the dispersed oil phase by the lipophilic radical generator, 2,2'-azobis(2,4-dimethylvaleronitrile). The extent of lipid oxidation is measured spectroscopically by following the loss of an oxidatively labile, lipophilic probe (methyl eleostearate), the synthesis of which is described here. Using this procedure, the ability of various aqueous phase solvated antioxidants (ascorbic acid, gallic acid, (-)-epicatechin, (-)-epigallocatechin-3-gallate) to regenerate lipid phase solvated α-tocopherol was evaluated. In all cases, the test compounds were able to inhibit oxidation reactions; however, these effects were not profoundly synergistic, and the maximum synergistic interaction observed was only ∼ 3% using ascorbic acid.

Topics: alpha-Tocopherol; Antioxidants; Ascorbic Acid; Catechin; Emulsions; Food; Gallic Acid; Humans; Hydrophobic and Hydrophilic Interactions; Linolenic Acids; Lipid Peroxidation; Lipids; Oxidation-Reduction

Malignant mesothelioma (MMe) is a poor-prognosis tumor in need of innovative therapies. In a previous in vivo study, we showed synergistic anti-MMe properties of the ascorbate/epigallocatechin-3-gallate/gemcitabine combination. We have now focused on the mechanism of action, showing the induction of apoptosis and cell cycle arrest through measurements of caspase 3, intracellular Ca(2+), annexin V, and DNA content. StellArray™ PCR technology and Western immunoblotting revealed DAPK2-dependent apoptosis, upregulation of cell cycle promoters, downregulation of cell cycle checkpoints and repression of NFκB expression. The complex of data indicates that the mixture is synergistic in inducing cell cycle deregulation and non-inflammatory apoptosis, suggesting its possible use in MMe treatment.

Topics: Apoptosis; Ascorbic Acid; Catechin; Cell Cycle; Cell Line, Tumor; Deoxycytidine; Drug Combinations; Drug Synergism; Gemcitabine; Humans; Mesothelioma

Epigallocatechin-gallate (EGCG) claims a plethora of health benefits including protection against neoplastic diseases. Meanwhile, heparan-sulfate proteoglycans (HSPGs) have defensive role against tumour cell invasion. Therefore, the chemopreventive and hepatoprotective effects of EGCG were studied in hepatocellular carcinoma (HCC) in vivo and in vitro and compared with strong water soluble antioxidant, sodium ascorbate.. HCC was induced in SD rats by thioacetamide (200 mg/Kg). Some rats were treated with EGCG (20 mg/Kg) or sodium ascorbate (100 mg/Kg). Liver impairment was assessed by measuring serum α-fetoprotein and investigating liver sections stained with H/E. Hepatic HSPGs, syndecan-1 and matrix metalloproteinase-9 (MMP-9) were measured by ELISA. Gene expression of fibroblast growth factor (FGF)-2 was measured. Cell death was assessed by caspase-3 activity. In addition, all markers were measured in human hepatocellular carcinoma cell line (HepG2).. EGCG increased the animal survival and decreased both α-fetoprotein and HepG2 viability. In addition, EGCG ameliorated fibrosis and massive hepatic tissue breakdown. EGCG restored HSPGs and reduced expression of MMP-9, syndecan-1 and FGF-2 in-vivo and in-vitro. Sodium ascorbate showed significantly lower results than EGCG.. Besides antioxidant activity, other mechanisms are involved in the chemopreventive and hepatoprotective effects of EGCG including restoration of HSPGs receptors and inhibition of vascular invasion.

Topics: alpha-Fetoproteins; Animals; Antineoplastic Agents, Phytogenic; Antioxidants; Ascorbic Acid; Camellia sinensis; Carcinoma, Hepatocellular; Catechin; Fibroblast Growth Factor 2; Gene Expression; Hep G2 Cells; Heparan Sulfate Proteoglycans; Humans; Liver; Liver Neoplasms; Male; Matrix Metalloproteinase 9; Neoplasm Invasiveness; Phytotherapy; Plant Extracts; Rats, Sprague-Dawley; Syndecan-1; Thioacetamide

Epigallocatechin gallate (EGCG) is a green tea catechin with potential health benefits, such as anti-oxidant, anti-carcinogenic and anti-inflammatory effects. In general, EGCG is highly susceptible to degradation, therefore presenting stability problems. The present paper was focused on the study of EGCG stability in HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) medium regarding the pH dependency, storage temperature and in the presence of ascorbic acid a reducing agent. The evaluation of EGCG in HEPES buffer has demonstrated that this molecule is not able of maintaining its physicochemical properties and potential beneficial effects, since it is partially or completely degraded, depending on the EGCG concentration. The storage temperature of EGCG most suitable to maintain its structure was shown to be the lower values (4 or -20 °C). The pH 3.5 was able to provide greater stability than pH 7.4. However, the presence of a reducing agent (i.e., ascorbic acid) was shown to provide greater protection against degradation of EGCG. A validation method based on RP-HPLC with UV-vis detection was carried out for two media: water and a biocompatible physiological medium composed of Transcutol®P, ethanol and ascorbic acid. The quantification of EGCG for purposes, using pure EGCG, requires a validated HPLC method which could be possible to apply in pharmacokinetic and pharmacodynamics studies.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Anticarcinogenic Agents; Antioxidants; Ascorbic Acid; Buffers; Catechin; Chemistry, Pharmaceutical; Chromatography, High Pressure Liquid; Chromatography, Reverse-Phase; Drug Stability; Drug Storage; Ethylene Glycols; Excipients; HEPES; Hot Temperature; Hydrogen-Ion Concentration; Oxidation-Reduction; Reducing Agents; Solvents; Spectrophotometry

Malignant pleural mesothelioma (MPM) is a poor prognosis disease lacking adequate therapy. We have previously shown that ascorbic acid administration is toxic to MPM cells. Here we evaluated a new combined therapy consisting of ascorbate/epigallocatechin-3-gallate/gemcitabine mixture (called AND, for Active Nutrients/Drug). In vitro effects of AND therapy on various MPM cell lines revealed a synergistic cytotoxic mechanism. In vivo experiments on a xenograft mouse model for MPM, obtained by REN cells injection in immunocompromised mice, showed that AND strongly reduced the size of primary tumor as well as the number and size of metastases, and prevented abdominal hemorrhage. Kaplan Meier curves and the log-rank test indicated a marked increase in the survival of AND-treated animals. Histochemical analysis of dissected tumors showed that AND induced a shift from cell proliferation to apoptosis in cancer cells. Lysates of tumors from AND-treated mice, analyzed with an antibody array, revealed decreased TIMP-1 and -2 expressions and no effects on angiogenesis regulating factors. Multiplex analysis for signaling protein phosphorylation exhibited inactivation of cell proliferation pathways. The complex of data showed that the AND treatment is synergistic in vitro on MPM cells, and blocks in vivo tumor progression and metastasization in REN-based xenografts. Hence, the AND combination is proposed as a new treatment for MPM.

Topics: Animals; Antineoplastic Combined Chemotherapy Protocols; Ascorbic Acid; Catechin; Cell Line, Tumor; Cell Proliferation; Deoxycytidine; Drug Synergism; Gemcitabine; Humans; Lung Neoplasms; Male; Mesothelioma; Mesothelioma, Malignant; Mice; Mice, Inbred NOD; Mice, SCID; Pleural Neoplasms; Signal Transduction; Xenograft Model Antitumor Assays

The effect of green tea formulated with vitamin C and xylitol on intestinal cell transport of gallated and nongallated catechin was studied. The transport of catechins from both apical to basolateral and basolateral to apical directions was measured. The effect of vitamin C (4, 10, 20 ppm), xylitol (11, 27.5, 55 ppm), and combinations of both on the intestinal transport rate of catechins was examined. The efflux value (Pb→a/Pa→b) of (-)-epigallocatechin (EGC), (-)-epigallocatechin gallate (EGCG), (-)-epicatechin (EC), and (-)-epicatechin gallate (ECG) was 0.26, 0.22, 1.22, and 0.17, respectively, indicating that EC appeared to be less absorbed compared with other catechins. The addition of xylitol (11, 27.5, 55 ppm) and vitamin C (4, 10, 20 ppm) and in combination enhanced transport rate of nongallated catechins such as EC and EGC. For EC, vitamin C was revealed to be the most effective on intestinal transport, implying the inhibition of the efflux transport mechanism of EC. Intestinal transport of gallated catechins significantly increased from catechins formulated with vitamin C and xylitol in a dose-dependent manner compared to the catechin-only formulation. Results provide a potential strategy to enhance the delivery and bioavailability of catechins in humans by modulating green tea formulation with vitamin C and xylitol.

Topics: Antioxidants; Ascorbic Acid; Biological Availability; Caco-2 Cells; Catechin; Chemistry, Pharmaceutical; Chromatography, High Pressure Liquid; Humans; Intestinal Absorption; Intestinal Mucosa; Intestines; Mass Spectrometry; Plant Extracts; Tea; Xylitol

Metastasis by cancer cells relies upon the acquisition of the ability to evade anoikis, a cell death process elicited by detachment from extracellular matrix (ECM). The molecular mechanisms that ECM-detached cancer cells use to survive are not understood. Striking increases in reactive oxygen species (ROS) occur in ECM-detached mammary epithelial cells, threatening cell viability by inhibiting ATP production, suggesting that ROS must be neutralized if cells are to survive ECM-detachment. Here, we report the discovery of a prominent role for antioxidant enzymes, including catalase and superoxide dismutase, in facilitating the survival of breast cancer cells after ECM-detachment. Enhanced expression of antioxidant enzymes in nonmalignant mammary epithelial cells detached from ECM resulted in ATP elevation and survival in the luminal space of mammary acini. Conversely, silencing antioxidant enzyme expression in multiple breast cancer cell lines caused ATP reduction and compromised anchorage-independent growth. Notably, antioxidant enzyme-deficient cancer cells were compromised in their ability to form tumors in mice. In aggregate, our results reveal a vital role for antioxidant enzyme activity in maintaining metabolic activity and anchorage-independent growth in breast cancer cells. Furthermore, these findings imply that eliminating antioxidant enzyme activity may be an effective strategy to enhance susceptibility to cell death in cancer cells that may otherwise survive ECM-detachment.

Topics: Adenosine Triphosphate; Animals; Antioxidants; Ascorbic Acid; Blotting, Western; Breast Neoplasms; Catalase; Catechin; Cell Adhesion; Cell Line; Cell Line, Tumor; Cell Survival; Chromans; Extracellular Matrix; Female; Humans; Mice; Mice, Nude; Reactive Oxygen Species; RNA Interference; Superoxide Dismutase; Tomography, X-Ray Computed; Xenograft Model Antitumor Assays

Tea polyphenols (-)-epigallocatechin-3-gallate (EGCG) and theaflavin-3-3'-digallate (TF3) are two prospective compounds in cancer prevention and treatment. Ascorbic acid (Vc) is essential to a healthy diet as well as being a highly effective antioxidant. In this work, the effects of the combination of EGCG or TF3 with Vc on the apoptosis and caspases-3/9 activities in human lung adenocarcinoma SPC-A-1 cells and esophageal carcinoma Eca-109 cells were determined. Furthermore, the role of mitogen-activated protein kinases (MAPK) pathways in the apoptosis induced by TF3 or EGCG together with Vc were studied using three MAPK inhibitors (ERK inhibitor PD98059, JNK inhibitor SP600125 and p38 inhibitor SB203580). Our results showed that Vc could enhance the EGCG and TF3 induced apoptosis in SPC-A-1 and Eca-109 cells, and this effect involved the activation of caspase-3 and 9. EGCG, TF3 and Vc could activate MAPK pathways respectively, and each compound activated different MAPK subfamilies in different cells. This may explain the enhancement of EGCG and TF3 induced apoptosis by Vc in SPC-A-1 and Eca-109 cells, and will ultimately aid the design of more effective anti-cancer treatments.

Topics: Adenocarcinoma; Anticarcinogenic Agents; Antineoplastic Agents; Apoptosis; Ascorbic Acid; Biflavonoids; Caspase 3; Caspase 9; Catechin; Cell Line, Tumor; Enzyme Inhibitors; Esophageal Neoplasms; Gallic Acid; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Humans; Lung Neoplasms; MAP Kinase Signaling System

The catechin (-)-epigallocatechin-3-gallate (EGCG) exhibits high antioxidant activity and it has been reported to provide protection of the skin against damage induced by solar UV radiation. However, EGCG is highly unstable under sunlight. The present study aimed to compare the effectiveness of the co-antioxidant agents vitamin E, butylated hydroxytoluene, vitamin C and a-lipoic acid for their potential to protect the catechin from photochemical degradation. Model creams (oil-in-water emulsions) containing EGCG (1%, w/w) alone or combined with equimolar concentrations of co-antioxidant were exposed to a solar simulator at an irradiance corresponding to natural sunlight. Photodegradation was evaluated by HPLC-UV and HPLC-ESI-MS/MS. Addition of the co-antioxidants vitamin C and a-lipoic acid to the formulation significantly reduced the light-induced decomposition of EGCG from 76.9 ± 4.6% to 20.4 ± 2.7% and 12.6 ± 1.6%, respectively. Conversely, butylated hydroxytoluene had no effect (EGCG loss, 78.1 ± 4.6%) and vitamin E enhanced the EGCG photolysis to 84.5 ± 3.4%. The functional stability of the catechin in the creams exposed to the solar simulator was also evaluated by measuring the in vitro antioxidant activity. Following irradiation, the reduction of the EGCG formulation antioxidant power was lower (21.8%) than the extent of degradation (76.9%), suggesting the formation of photoproducts with antioxidant properties. The influence of the examined co-antioxidants on the functional stability of the catechin under simulated sunlight paralleled that measured for the EGCG photodecomposition, a-lipoic acid exerting the greatest stabilising effect (antioxidant activity decrease, 1.4%). These results demonstrated that a-lipoic acid is an effective co-antioxidant agent for the stabilization of EGCG in dermatological products for skin photoprotection.

Topics: Antioxidants; Ascorbic Acid; Biphenyl Compounds; Butylated Hydroxytoluene; Catechin; Chemistry, Pharmaceutical; Chromatography, High Pressure Liquid; Drug Stability; Emulsions; Free Radicals; Light; Oxidation-Reduction; Photolysis; Picrates; Skin Cream; Spectrometry, Mass, Electrospray Ionization; Tandem Mass Spectrometry; Thioctic Acid; Vitamin E

5-Caffeoylquinic acid (5-CQA) is generally referred to as chlorogenic acid and exhibits various biological activities such as antioxidant activity and porcine pancreas α-amylase inhibitory activities. 5-CQA may be useful as an antioxidant for food and to prevent diabetes and obesity. The degradation of 5-CQA and caffeic acid (CA) in an aqueous solution at 37 °C and pH 5.0-9.0 was studied. The degradation of 5-CQA and CA, demonstrating time and pH dependence (i.e., the rate constant, k, was higher at higher pH), was satisfactorily described by the Weibull equation. The stability of 5-CQA at pH 7.4 and 9.0 was improved by adding (-)-epigallocatechin gallate (EGCG) and ascorbic acid (AA). Moreover, the degradation of 5-CQA in the presence of EGCG or AA could be described by the Weibull equation. The k value in the presence of EGCG or AA was dependent on their concentration.

Topics: Ascorbic Acid; Caffeic Acids; Catechin; Chlorogenic Acid; Drug Stability; Hydrogen-Ion Concentration; Kinetics; Quinic Acid

The mechanism of iron release from ferritin in vivo is still unclear even though it represents a key step of the metabolism of iron in vivo. Here, both interaction intensity and binding stability between epigallocatechin gallate (EGCG) from tea and liver ferritin of Dasyatis akajei (DALF) were investigated using UV-visible, fluorescence and circular dichroism (CD) spectrometry, respectively. The results indicated that EGCG could reduce the iron within the ferritin shell directly in the absence of chemical reducers such as Na(2)S(2)O(4), but this process was strictly pH-dependent, and the rate of iron release is faster at low pH than at high pH. The kinetic study of iron release showed that this process fitted the law of zero order reaction, which differed from that of first order reaction by various chemical reducers such as Vitamin C. In addition, Both fluorescence and CD spectrometry were further used to study the reduction mechanism of iron release in vitro, showing that there was a slight conformation change of the ferritin shell during EGCG reduction because of a complex formation of DALF-EGCG. It appears that chemical reducers with large molecular sizes reduce the iron across the protein shell by the way of an electron transfer pathway (ETP). A novel pathway for iron release from DALF with EGCG reduction is suggested to explain for a reductive route of iron metabolism by biological reducers in vivo.

Topics: Animals; Ascorbic Acid; Catechin; Circular Dichroism; Elasmobranchii; Ferritins; Hydrogen-Ion Concentration; Iron; Kinetics; Liver

Several phenolic compounds as well as ascorbate can oxidise in certain cell culture media (especially Dulbecco's modified Eagle's medium (DMEM)) to generate hydrogen peroxide. Addition of oxaloacetate decreased the levels of H(2)O(2) detected and the oxaloacetate was depleted. Oxaloacetate was approximately as effective as pyruvate in decreasing H(2)O(2) levels and more effective than α-ketoglutarate. Our data raise important issues to consider when interpreting the behaviour and metabolism of cells in culture (which are both altered by the oxidative stress of cell culture) and their apparent response to addition of autooxidisable compounds such as ascorbate and epigallocatechin gallate.

Topics: Ascorbic Acid; Catechin; Cells; Cells, Cultured; Culture Media; Hydrogen Peroxide; Oxaloacetic Acid

While flavonoids can reportedly protect against cardiac ischemia-reperfusion injury, the relative effectiveness of different flavonoids and the mechanisms involved are unclear. We compared protection by different flavonoids using rat embryonic ventricular H9c2 cells subjected to simulated ischemia-reperfusion (IR) and to tert-butyl hydroperoxide (t-buOOH). Characterization of the IR model showed the relative contributions of glucose, serum, and oxygen deprivation to cell death. With long-term (2-3 day) pretreatment before IR the best protection was given by catechin, epigallocatechin gallate, proanthocyanidins, and ascorbate, which protected at all doses. Quercetin protected (34%) at 5 μM but was cytotoxic at higher doses. Cyanidin protected mildly (10-15%) at 5 and 20 μM, while delphinidin had no effect at 5 μM and was cytotoxic at higher doses. Comparing long-term and acute protection by catechin, a higher concentration was needed for benefit with acute (1 hr) pretreatment. With a pure oxidative stress (t-buOOH) only quercetin significantly protected with 3-day pretreatment, while with short-term (1 h) pretreatments protection was best with quercetin and epigallocatechin gallate. The results suggest catechins to be especially useful as IR preconditioning agents, while quercetin and epigallocatechin gallate may be the most protective acutely in situations of oxidative stress.

Topics: Animals; Ascorbic Acid; Cardiotonic Agents; Catechin; Cell Death; Cell Line; Culture Media, Serum-Free; Cytoprotection; Flavonoids; Glucose; Glyceraldehyde-3-Phosphate Dehydrogenases; Myocardial Reperfusion Injury; Myocytes, Cardiac; Oxidation-Reduction; Oxidative Stress; Oxygen; Quercetin; Rats; tert-Butylhydroperoxide; Time Factors

Increasing evidence has demonstrated that EGCG possesses prooxidant potential in biological systems, including modifying proteins, breaking DNA strands and inducing the generation of reactive oxygen species. In the present study, the prooxidant effect of EGCG on erythrocyte membranes was investigated. SDS-PAGE and NBT-staining assay were utilized to detect the catechol-protein adducts that generated upon treating the membranes with EGCG. The results indicated that EGCG was able to bind covalently to sulfhydryl groups of membrane proteins, leading to the formation of protein aggregates with intermolecular cross-linking. We suggested that the catechol-quinone originated from the oxidation of EGCG acted as a cross-linker on which peptide chains were combined through thiol-S-alkylation at the C2- and C6-sites of the gallyl ring. EGC showed similar effects as EGCG on the ghost membranes, whereas ECG and EC did not, suggesting that a structure with a gallyl moiety is a prerequisite for a catechin to induce the aggregation of membrane proteins and to deplete membrane sulfhydryls. EDTA and ascorbic acid inhibited the EGCG-induced aggregation of membrane proteins by blocking the formation of catechol-quinone. The information of the present study may provide a fresh insight into the prooxidant effect and cytotoxicity of tea catechins.

Topics: Ascorbic Acid; Benzoquinones; Catechin; Cell Membrane; Cross-Linking Reagents; Edetic Acid; Erythrocytes; Flavonoids; Humans; Membrane Proteins; Molecular Weight; Phenols; Polyphenols; Protein Multimerization; Protein Structure, Quaternary; Reactive Oxygen Species; Sulfhydryl Compounds; Tea

Ascorbate and several phenolic compounds readily oxidise in cell culture media to generate hydrogen peroxide. However, addition of α-ketoglutarate, which is known to be released by several cell types, decreased the levels of H(2)O(2), and the α-ketoglutarate was depleted and converted to succinate. These observations could account for previous reports of the protective effects of α-ketoglutarate in promoting the growth of cells in culture, and may contribute to explaining some of the variability in the literature in reported rates of H(2)O(2) production from autoxidisable compounds in cell culture systems.

Topics: Animals; Artifacts; Ascorbic Acid; Catechin; Cell Culture Techniques; Cells, Cultured; Culture Media; Hydrogen Peroxide; Ketoglutaric Acids; Oxidation-Reduction; Succinic Acid

To investigate whether clinically relevant levels of epigallocatechin gallate (EGCG, a component of green tea) or vitamin C (ascorbic acid) could antagonize bortezomib antitumor activity in CWR22 human prostate xenograft tumors.. The pharmacokinetics (PK) of EGCG and ascorbic acid were determined in immunocompromised mice and compared with concentrations measured in human PK studies of dietary supplements. Antitumor activity of bortezomib in combination with EGCG or ascorbic acid was determined using several dosing regimens to evaluate different target plasma concentrations of EGCG and ascorbic acid.. Bortezomib dosed twice-weekly at 0.8 mg/kg IV demonstrated tumor growth inhibition (TGI) of 53.9-58.9%. However, when combined with EGCG such that the plasma concentrations of EGCG were >200 μM at the time of bortezomib dosing, all antitumor activity was abrogated (TGI = -17.7%). A lower concentration of EGCG (11-16 μM), which is severalfold higher than measured clinically in humans taking EGCG supplements (0.6-3 μM), was not antagonistic to bortezomib (TGI 63.5%). Pharmacodynamic studies of proteasome inhibition reflected these findings. Ascorbic acid (40 and 500 mg/kg PO daily) was evaluated under a similar study design and did not antagonize bortezomib antitumor activity (TGI 57.2 and 72.2%).. No antagonism of bortezomib is seen in preclinical in vivo experiments, where EGCG or ascorbic acid plasma concentrations are commensurate with dietary or supplemental intake. The data suggest that patients receiving bortezomib treatment do not need to avoid normal dietary consumption of green tea, vitamin C-containing foods, or EGCG or vitamin C dietary supplements.

Topics: Animals; Antineoplastic Agents; Ascorbic Acid; Boronic Acids; Bortezomib; Catechin; Chromatography, Liquid; Drug Screening Assays, Antitumor; Female; Humans; Male; Mice; Mice, SCID; Prostatic Neoplasms; Pyrazines; Tandem Mass Spectrometry; Tea; Xenograft Model Antitumor Assays

We previously reported that (-)-epigallocatechin-3-gallate (EGCG) and grape seed extract (GSE) at high concentration nearly blocked intestinal iron transport across the enterocyte. In this study, we aimed to determine whether small amounts of EGCG, GSE, and green tea extract (GT) are capable of inhibiting iron absorption, to examine if ascorbic acid counteracts the inhibitory action of polyphenols on iron absorption, and to explore the mechanisms of polyphenol-mediated apical iron uptake and basolateral iron release. An(55)Fe absorption study was conducted by adding various concentrations of EGCG, GSE, and GT using Caco-2 intestinal cells. Polyphenols were found to inhibit the transepithelial (55)Fe transport in a dose-dependent manner. The addition of ascorbic acid offset the inhibitory effects of polyphenols on iron transport. Ascorbic acid modulated the transepithelial iron transport without changing the apical iron uptake and the expression of ferroportin-1 protein in the presence of EGCG. The polyphenol-mediated apical iron uptake was inhibited by membrane impermeable Fe(2+) chelators (P < 0.001), but at a low temperature (4°C), the apical iron uptake was still higher than the control values at 37°C (P < 0.001). These results suggest that polyphenols enhance the apical iron uptake partially by reducing the conversion of ferric to ferrous ions and possibly by increasing the uptake of polyphenol-iron complexes via the energy-independent pathway. The present results indicate that the inhibitory effects of dietary polyphenols on iron absorption can be offset by ascorbic acid. Further studies are needed to confirm the current findings in vivo.

Topics: Ascorbic Acid; Biological Transport; Caco-2 Cells; Catechin; Cation Transport Proteins; Cell Polarity; Cold Temperature; Diet; Dietary Supplements; Enterocytes; Flavonoids; Grape Seed Extract; Humans; Intestinal Absorption; Iron Chelating Agents; Iron Radioisotopes; Iron, Dietary; Oxidation-Reduction; Phenols; Plant Extracts; Polyphenols; Tea

Malignant mesothelioma (MMe) is a lethal tumor arising from the mesothelium of serous cavities as a result of exposure to asbestos. Current clinical standards consist of combined treatments, but an effective therapy has not been established yet and there is an urgent need for new curative approaches. Ascorbate is a nutrient that is also known as a remedy in the treatment of cancer. In the present study, we have tested the cytotoxicity of ascorbate to MMe cells in combination with drugs used in MMe therapy, such as cisplatin, etoposide, gemcitabine, imatinib, paclitaxel, and raltitrexed, as well as with promising antitumor compounds like taurolidine, α-tocopherol succinate, and epigallocatechin-3-gallate (EGCG). Dose-response curves obtained for each compound by applying the neutral red uptake (NRU) assay to MMe cells growing in vitro, allowed to obtain IC50 values for each compound used singularly. Thereafter, NRU data obtained from each ascorbate/drug combination were analyzed through Tallarida's isobolograms at the IC50 level (Tallarida, 2000), revealing synergistic interactions for ascorbate/gemcitabine and ascorbate/EGCG. These results were further confirmed through comparisons between theoretical additivity IC50 and observed IC50 from fixed-ratio dose-response curves, and over a broad range of IC levels, by using Chou and Talalay's combination index (Chou and Talalay, 1984). Synergistic interactions were also shown by examining apoptosis and necrosis rates, using the caspase 3 and lactic dehydrogenase assays, respectively. Hence, data indicate that ascorbate/gemcitabine and ascorbate/EGCG affect synergistically the viability of MMe cells and suggest their possible use in the clinical treatment of this problematic cancer.

Topics: Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Ascorbic Acid; Catechin; Cell Line, Tumor; Cell Proliferation; Cell Survival; Coloring Agents; Deoxycytidine; Drug Synergism; Gemcitabine; Humans; L-Lactate Dehydrogenase; Lung Neoplasms; Mesothelioma; Neutral Red

Chromatographic separation of black tea polyphenols is too difficult to supply sufficient quantities of pure compounds for biological experiments. Thus, facile methods to prepare black tea constituents were desired. Treatment of epigallocatechin gallate with copper(II) chloride efficiently afforded an unstable quinone dimer, dehydrotheasinensin A, and subsequent treatment with ascorbic acid stereoselectively yielded theasinensin A. The latter is a dimer with an R-biphenyl bond, one of the major polyphenols found in black tea. The method is simpler and more effective than enzymatic preparation.

Topics: Ascorbic Acid; Benzopyrans; Biomimetic Materials; Catechin; Copper; Dimerization; Oxidation-Reduction; Phenols; Polyphenols; Stereoisomerism; Tea

With a view of simultaneous determination of physio-active ingredients in oolong tea infusion: sugars, amino acids, epigallocatechin gallate and ascorbic acid, a novel CZE with amperometric detection method was studied. Operated in a wall-jet configuration, 100 mmol/L NaOH was used in detecting cell to lead the electrocatalysis oxidation behaviors of the analytes on a 300 mum diameter copper-disc electrode (working electrode), while in separating capillary, a mild alkaline running buffer consisting in a mixture of 30 mmol/L borate and 40 mmol/L phosphates charged and carried analytes to detecting end. The methodology research was performed for system stability and suitability. Under the optimal CE conditions, analytes could be separated within moderate time period. Good linearity between peak area and concentration existed over three orders of magnitude; lower RSD and LOD were achieved. The oolong tea infusion was assayed and result was satisfactory.

Topics: Amino Acids; Ascorbic Acid; Buffers; Carbohydrates; Catechin; Electrochemistry; Electrophoresis, Capillary; Limit of Detection; Reproducibility of Results; Tea

Oxidation of polyunsaturated fatty acids (PUFAs) releases alpha,beta-unsaturated aldehydes that modify deoxyguanosine (dG) to form cyclic 1,N(2)-propanodeoxyguanosine adducts. One of the major adducts detected in vivo is acrolein (Acr)-derived 1,N(2)-propanodeoxyguanosine (Acr-dG). We used a chemical model system to examine the effects of 4 antioxidants known to inhibit fatty acid oxidation on the formation of Acr-dG and 8-oxodeoxyguaonsine (8-oxodG) from the PUFA docosahexaenoic acid (DHA) under oxidative conditions. We found that epigallocatechin gallate (EGCG) and dihydrolipoic acid (DHLA) inhibit both Acr-dG and 8-oxodG formation. In contrast, ascorbic acid and alpha-tocopherol actually increase Acr-dG at high concentrations and do not show a concentration-dependant inhibition of 8-oxodG. We also studied their effects on blocking Acr-dG formation directly from Acr. EGCG and DHLA can both effectively block Acr-dG formation, but ascorbic acid and alpha-tocopherol show weak or little effect. These results highlight the complexity of antioxidant mechanisms and also reveal that EGCG and DHLA are effective at suppressing lipid peroxidation-induced Acr-dG and 8-oxodG formation as well as blocking the reaction of dG with Acr.

Topics: 8-Hydroxy-2'-Deoxyguanosine; Acrolein; alpha-Tocopherol; Antioxidants; Ascorbic Acid; Catechin; Deoxyguanosine; DNA Damage; Docosahexaenoic Acids; Lipid Peroxidation; Thioctic Acid

Type IV collagenase matrix metalloproteinases (MMPs), especially MMP-2 and MMP-9, have been found to promote invasion and metastasis of malignant tumors. Extracellular matrix (ECM) degradation by MMPs and increased expression of MMPs in cancer cells and tumor microvascular endothelial cells make MMPs an attractive target for cancer. Focused on a common pathomechanism of cancer growth and invasion, the disintegration of connective tissue, we used natural approaches to increase the integrity and strength of connective tissues. Utilizing the principle of nutrition synergy, we developed a novel micronutrient mixture (NM) containing lysine, proline, ascorbic acid and green tea extract. This study evaluates the potency of the components EGCG and green tea extract independently compared to that of NM on modulation of patterns of MMP-2 and MMP-9 expression in four cancer cell lines expressing MMP-2, MMP-9 or both. Human fibrosarcoma (HT-1080), hepatocellular carcinoma (SK-Hep-1), glioblastoma (T-98G), uterine leiomyosarcoma (SK-UT-1) cell lines were obtained from ATCC and grown in minimum essential medium (MEM) supplemented with 10% FBS, penicillin (100 U/ml) and streptomycin (100 mg/ml) in 24-well tissue culture plates. At near confluence, the cells were treated with agents dissolved in media and tested at concentrations indicated in triplicate at each dose. Cells were also treated with PMA 100 ng/ml to study enhanced expression of MMP-9. MMP expression was assessed by gelatinase zymography. Fibrosarcoma and hepatocellular carcinoma cells expressed both MMP-2 and MMP-9. Glioblastoma cells expressed MMP-2 and PMA treatment induced MMP-9 expression. Uterine leimyosarcoma cells expressed no MMPs but PMA induced MMP-9. NM was the most potent dose-dependent inhibitor of MMPs, followed by green tea extract and EGCG. In conclusion, these results suggest the enhanced efficacy of nutrients working in synergy to modulate complex pathways such as MMP expression.

Topics: Antineoplastic Agents, Phytogenic; Ascorbic Acid; Camellia sinensis; Catechin; Cell Line, Tumor; Dose-Response Relationship, Drug; Humans; Lysine; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Micronutrients; Neoplasm Invasiveness; Neoplasms; Plant Extracts; Proline; Protease Inhibitors; Tetradecanoylphorbol Acetate

Dietary supplements have been extensively studied for their beneficial effects on cognition and AD neuropathology. The current study examines the effect of a medical food cocktail consisting of the dietary supplements curcumin, piperine, epigallocatechin gallate, α-lipoic acid, N-acetylcysteine, B vitamins, vitamin C, and folate on cognitive functioning and the AD hallmark features and amyloid-beta (Aβ) in the Tg2576 mouse model of the disease.. The study found that administering the medical food cocktail for 6 months improved cortical- and hippocampal- dependent learning in the transgenic mice, rendering their performance indistinguishable from non-transgenic controls. Coinciding with this improvement in learning and memory, we found that treatment resulted in decreased soluble Aβ, including Aβ oligomers, previously found to be linked to cognitive functioning.. In conclusion, the current study demonstrates that combination diet consisting of natural dietary supplements improves cognitive functioning while decreasing AD neuropathology and may thus represent a safe, natural treatment for AD.

Topics: Acetylcysteine; Alkaloids; Alzheimer Disease; Amyloid beta-Protein Precursor; Animals; Ascorbic Acid; Benzodioxoles; Brain; Catechin; Cerebral Cortex; Cognition; Curcumin; Dietary Supplements; Folic Acid; Hippocampus; Humans; Immunoblotting; Maze Learning; Memory; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Transgenic; Piperidines; Polyunsaturated Alkamides; Thioctic Acid; Vitamin B Complex; Vitamins

Ascorbate and several phenolic compounds readily oxidise in cell culture media to generate hydrogen peroxide. However, media containing pyruvate showed much less H(2)O(2) production, apparently because pyruvate can scavenge H(2)O(2) in the medium. Researchers must be aware that compounds under test can sometimes readily oxidise in cell culture media, that this might not be detected by measurement of H(2)O(2) if the media contain pyruvate, and that pyruvate can be substantially depleted in the media as a result.

Topics: Artifacts; Ascorbic Acid; Catechin; Culture Media; Free Radical Scavengers; Hydrogen Peroxide; Pyruvic Acid

In mammals, aging is linked to a decline in the activity of citrate synthase (CS; E.C. 2.3.3.1), the first enzyme of the citric acid cycle. We used 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH), a water-soluble generator of peroxyl and alkoxyl radicals, to investigate the susceptibility of CS to oxidative damage. Treatment of isolated mitochondria with AAPH for 8-24 h led to CS inactivation; however, the activity of aconitase, a mitochondrial enzyme routinely used as an oxidative stress marker, was unaffected. In addition to enzyme inactivation, AAPH treatment of purified CS resulted in dityrosine formation, increased protein surface hydrophobicity, and loss of tryptophan fluorescence. Propyl gallate, 1,8-naphthalenediol, 2,3-naphthalenediol, ascorbic acid, glutathione, and oxaloacetate protected CS from AAPH-mediated inactivation, with IC(50) values of 9, 14, 34, 37, 150, and 160 muM, respectively. Surprisingly, the antioxidant epigallocatechin gallate offered no protection against AAPH, but instead caused CS inactivation. Our results suggest that the current practice of using the enzymatic activity of CS as an index of mitochondrial abundance and the use of aconitase activity as an oxidative stress marker may be inappropriate, especially in oxidative stress-related studies, during which alkyl peroxyl and alkoxyl radicals can be generated.

Topics: Aconitate Hydratase; Amidines; Antioxidants; Ascorbic Acid; Catechin; Citrate (si)-Synthase; Enzyme Activation; Glutathione; Inhibitory Concentration 50; Mitochondria; Naphthols; Oxaloacetic Acid; Oxidants; Oxidation-Reduction; Oxidative Stress; Peroxides; Propyl Gallate

Oxidative stress resulting from an imbalance between radical-generating and radical scavenging systems plays an important role in the pathogenesis of pulmonary fibrosis. Epigallocatechin-3-gallate (EGCG), a polyphenol and a major component of green tea, possess a potent antioxidant property. This study was designed to evaluate the potential antioxidative activity of EGCG in the plasma and lungs during bleomycin induced experimental pulmonary fibrosis. Intratracheal administration of bleomycin (6.5 U/kg body weight) to rats resulted in significant reduction of body weight, enzymic antioxidants (superoxide dismutase, catalase, glutathione peroxidase and glutathione reductase) and non-enzymic antioxidants (reduced glutathione, vitamin C, vitamin E and vitamin A). Elevations in lung W/D (wet weight/dry weight) ratio, hydroxyproline content was observed with a synchronized increase in lipid peroxidation markers (thiobarbituric acid reactive substances and hydroperoxides). Intraperitoneal administration of EGCG at a dose of 20 mg/kg body weight significantly improved the body weight, enzymic and non enzymic antioxidants and considerably decreased the W/D ratio, hydroxyproline and lipid peroxidation marker levels. Histological observations also correlated with the biochemical parameters. Thus, this study confirms the beneficial use of EGCG in alleviating the oxidative stress induced during pulmonary fibrosis.

Topics: Animals; Antibiotics, Antineoplastic; Antioxidants; Ascorbic Acid; Bleomycin; Body Weight; Catechin; Glutathione; Injections, Intraperitoneal; Lung; Male; Organ Size; Pulmonary Fibrosis; Rats; Rats, Wistar; Thiobarbituric Acid Reactive Substances; Vitamin E

The aim of the study was to establish a 96-well microtiter plate-based reporter gene assay to test the influence of natural compounds on the promoter activities of rat catalase, human glutathione peroxidase and human superoxide dismutase expressed in V79 cells. Luciferase expression vectors with the promoter regions of the genes coding for the three above-mentioned enzymes were constructed and transfected into V79 cells. Thereafter the ability of sodium ascorbate, L-carnitine, catechin, epigallocatechin gallate, genistein, paraquat, quercetin, 12-O-tetradecanoylphorbol-13-acetate and Trolox to enhance the promoter activities was evaluated. Genistein, paraquat and quercetin led to a statistically significant increase in the glutathione peroxidase and superoxide dismutase gene promoter activities. None of the compounds tested enhanced the catalase gene promoter activity. The reporter gene assay described in this report is easy to perform, fast and allows one to test a high number of compounds and different concentrations of a single compound at the same time.

Topics: Animals; Anticarcinogenic Agents; Antioxidants; Ascorbic Acid; Carcinogens; Carnitine; Catalase; Catechin; Cells, Cultured; Chromans; Cricetinae; Cricetulus; Genistein; Glutathione Peroxidase; Herbicides; Humans; Luciferases; Paraquat; Promoter Regions, Genetic; Quercetin; Rats; Superoxide Dismutase; Tetradecanoylphorbol Acetate; Vitamin B Complex

Although plant polyphenols such as (-)-epigallocatechin gallate (EGCG) have antibacterial activity towards methicillin-resistant Staphylococcus aureus (MRSA), such polyphenols are unstable in solution. Because the instability of polyphenols is attributable to their oxidation, we examined the effects of antioxidants and inhibitors of polyphenol oxidation on the maintenance of polyphenol antibacterial activity. The antibacterial activity of EGCG was enhanced in the presence of ascorbic acid, and ascorbic acid was the most effective for retaining the concentration of stable EGCG. On the other hand, the antibacterial activity of EGCG was lowered in the presence of casein in spite of its suppressing effect on the EGCG decrease. The effect of EGCG on the antibiotic resistance of MRSA was also enhanced in the presence of ascorbic acid. The addition of an antioxidant may affect other pharmacological effects of polyphenols in analogous ways, although this does not mean the clinical usefulness of the addition directly.

Topics: Anti-Bacterial Agents; Ascorbic Acid; Catechin; Methicillin-Resistant Staphylococcus aureus; Molecular Structure; Oxidation-Reduction; Time Factors

The effect of photosensitized oxidation of conjugated linoleic acid in an oil-in-water (o/w) emulsion system was studied. Water-soluble natural antioxidants, including apple polyphenols from apple extract, green tea extract, 4-hydroxy-2(or 5)-ethyl-5(or2)-methyl-3(2H)-furanone(HEMF), 4-hydroxy-2,5-dimethyl-3(2H)-furanone (HDMF), and ascorbic acid, were tested for antioxidant activity in this system. The green tea extract showed the highest antioxidant activity followed by ascorbic acid. Apple polyphenols did not give significant antioxidant activity. HEMF and HDMF exhibited a prooxidant effect. The antioxidant activity of tea catechins was also investigated. Of them, EGCG and ECG exhibited antioxidant activity at 50 ppm, but the antioxidant activity between them was not significantly different (P < 0.05). Comparatively, EC, EGC, and GCG showed no significant antioxidative effect at 50 ppm. When the concentration increased to 100 ppm, the antioxidant activity of ECG and EGCG significantly increased compared with that at 50 ppm, and EGCG had higher antioxidant activity than ECG. GCG also showed significant antioxidant activity at 100 ppm. EGCG exhibited the highest antioxidant activity among the tea catechins in the emulsion system at 100 ppm.

Topics: Antioxidants; Ascorbic Acid; Catechin; Emulsions; Flavonoids; Fruit; Furans; Light; Linoleic Acids, Conjugated; Oxidation-Reduction; Phenols; Polyphenols; Singlet Oxygen; Solubility; Vitis; Water

Oxidative stress is believed to contribute to the pathogenesis of hypercholesterolaemic atherosclerosis; hence, various antioxidant compounds are being evaluated for potential anti-atherogenic effects. The present study assessed the efficacy of epigallocatechin gallate (EGCG), an antioxidant component of the plant Camellia sinensis, in improving serum lipid profile and antioxidant parameters in erythrocytes and cardiac tissue in rats fed an atherogenic diet. In male albino Wistar rats fed an atherogenic diet for 30 days, significantly increased serum levels of total cholesterol, triglycerides and lipoprotein cholesterol fractions and cardiac risk ratio were noted, compared with levels in rats fed a normal diet. Intraperitoneal administration of EGCG (100 mg/kg) for 7 or 15 days to the atherogenic diet-fed rats resulted in significantly lower serum levels of total cholesterol, triglycerides, low-density and very low density lipoprotein cholesterol fractions and a significantly higher serum level of high-density lipoprotein cholesterol compared with levels in atherogenic diet-fed, saline-treated rats. Significantly higher mean malondialdehyde levels and significantly lower mean activities of antioxidant enzymes and mean levels of non-enzymatic antioxidants occurred in atherogenic diet-fed rats compared with those fed a normal diet. When atherogenic diet-fed rats received EGCG treatment for 7 or 15 days, significantly lower mean levels of MDA, higher mean levels of non-enzymatic antioxidants and higher mean activities of enzymatic antioxidants occurred, compared with those in saline-treated rats. Thus, EGCG appears to ameliorate disruptions of serum lipid profile and of antioxidant parameters in erythrocyte and cardiac tissue of Wistar rats fed an atherogenic diet; these results may be relevant to treating human atherosclerosis.

Topics: Animals; Anticholesteremic Agents; Antioxidants; Aorta, Thoracic; Ascorbic Acid; Atherosclerosis; Camellia sinensis; Catalase; Catechin; Diet, Atherogenic; Erythrocytes; Glutathione Peroxidase; Hypercholesterolemia; Injections, Intraperitoneal; Lipid Peroxidation; Lipids; Male; Oxidative Stress; Plant Extracts; Rats; Rats, Wistar; Superoxide Dismutase; Vitamin E

The aim of this study was to investigate the mechanism of antioxidant and prooxidant effects of epigallocatechin-3-gallate (EGCG) using chemiluminescence analysis. Results showed that EGCG scavenged superoxide radical and H2O22 in a dose dependent manner. EGCG scavenged 50% of superoxide radical at 0.31 mM and scavenged 50% of H2O22 at 0.09 mM, demonstrating that EGCG has a stronger reactive oxygen species (ROS) scavenging activity than ascorbic acid. Effects of EGCG on free radical-induced DNA oxidative damage were investigated. EGCG had protective effect on DNA at low concentrations (2-30 mM), but it enhanced the DNA oxidative damage at higher concentrations (>60 mM), exhibiting a prooxidant effect on DNA. EGCG showed a greater reducing power on iron ions, reducing Fe3+ to Fe2+, which accelerates the generation of hydroxyl radical from the Fenton reaction. At low concentrations, ROS scavenging activity of EGCG might predominate over its reducing power and lead to its protective effect on DNA. However, relatively higher reducing power of EGCG at higher concentrations may gradually predominate over its ROS scavenging activity and result in the prooxidant effect of EGCG on DNA.

Topics: Antioxidants; Ascorbic Acid; Catechin; DNA Damage; Dose-Response Relationship, Drug; Food, Organic; Humans; Hydrogen Peroxide; Iron; Luminescence; Oxidation-Reduction; Reactive Oxygen Species; Superoxides; Tea

Certain drastic behavioral modifications by arterial wall smooth muscle cells (SMC) have been considered key steps in the formation of atherosclerotic lesions: massive migration of SMC from the media to the intima layer of the vessel, dedifferentiation of SMC to proliferating phenotype, and increased secretion of inflammatory cytokines as a response to inflammatory stimuli. We investigated the anti-atherogenic effects of naturally occurring compounds (ascorbic acid, green tea extract, lysine, proline, arginine, and N-acetyl cysteine) using the model of cultured aortic SMC. Cell growth was measured by DNA synthesis, cell invasiveness was measured through Matrigel, matrix metalloproteinase-2 (MMP-2) secretion was measured by zymography, and SMC secretion of monocyte chemoattractant protein-1 (MCP-1) and interleukin-6 (IL-6) was measured by immunochemistry. Fetal bovine serum-stimulated SMC growth was inhibited by the nutrient mixture (NM) with 85% inhibition at 100 microg/mL. A corresponding concentration of epigallocatechin gallate (EGCG; 15 microM), the most active tea phenolic, produced a significant effect but one lower than NM. NM inhibited aortic SMC Matrigel invasion in a dose-dependent manner and significantly decreased MMP-2 expression. Stimulation of SMC with tumor necrosis factor-alpha significantly increased production and secretion of such mediators of inflammation as IL-6 and MCP-1; addition of 100 microg/mL NM inhibited secretion of MCP-1 and IL-6 by 65% and 47%, respectively. These data suggest that the NM of ascorbic acid, tea phenolics, and selected amino acids has potential in blocking the development of atherosclerotic lesions by inhibiting atherogenic responses of vascular SMC to pathologic stimuli and warrants in vivo studies.

Topics: Amino Acids; Animals; Aorta; Ascorbic Acid; Atherosclerosis; Catechin; Cattle; Cell Proliferation; Cells, Cultured; Chemokine CCL2; Dose-Response Relationship, Drug; Humans; Immunochemistry; Interleukin-6; Matrix Metalloproteinase 2; Myocytes, Smooth Muscle; Phenols; Plant Extracts; Tea

Oxidative stress has been linked to the development of various chronic diseases. Vegetables and fruits, which contain polyphenols, were shown to have protective effects. (-)-Epigallocatechin-3-gallate (EGCG), a polyphenol abundant in tea, has been shown to have antioxidant activities in cell-free conditions and this study focused on the effect of cellular EGCG. Using an intestinal cell model to examine the oxidative stress induced by hydroxyl radicals, we report here that physiological concentrations (0.1-1 microM) of EGCG have dose- and incubation duration-dependent cell-associated lipid antioxidant activity (measuring malondialdehyde production). Vitamin E and vitamin C at 10-40 microM also showed cell-associated lipid antioxidant activities under shorter incubation durations. When EGCG was included in the incubation with vitamin E or C, more antioxidant activities were consistently observed than when vitamins were added alone. Catechin (widely present in fruits and vegetables) at 1 microM also significantly increased the antioxidant activity of vitamins E and C. Previous studies examining cell-associated activity of EGCG mainly focused on the 10-100 microM concentration range. Our results suggest that although the physiological level (0.1-1 microM) of dietary catechins is much lower than that of vitamins, they further contribute to the total antioxidant capacity even in the presence of vitamins.

Topics: Antioxidants; Ascorbic Acid; Caco-2 Cells; Catechin; Humans; Intestinal Mucosa; Intestines; Lipid Metabolism; Malondialdehyde; Tea; Vitamin E

The effects of a novel nutrient formulation Epican Forte (EF) were evaluated on proliferation and induction of apoptosis using non-cytotoxic concentrations against HTLV-1 positive (HuT-102 & C91-PL) and negative (CEM & Jurkat) cells. EF showed anti-proliferative effect as determined by MTT assay and TGF mRNA protein expression using RT-PCR. EF resulted in the down-regulation of TGF-alpha and an up-regulation in TGF-beta2. EF caused a significant increase in apoptotic cells in the preG1 phase. These results were confirmed using Cell Death ELISA and Annexin V-FITC. Induction of apoptosis was caused by an up-regulation of p53, p21 and Bax protein levels and a down-regulation of Bcl-2alpha protein expression level.

Topics: Acetylcysteine; Antineoplastic Agents; Apoptosis; Arginine; Ascorbic Acid; Catechin; Cell Cycle; Cell Line, Tumor; Cell Proliferation; Copper; Dietary Supplements; DNA; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Gene Expression Profiling; Human T-lymphotropic virus 1; Humans; Lysine; Manganese; Proline; Reverse Transcriptase Polymerase Chain Reaction; Selenium; Sensitivity and Specificity; Structure-Activity Relationship; T-Lymphocytes; Transforming Growth Factors

Following oxidative stress, modifications of several biologically important macromolecules have been demonstrated. In this study we investigated the effect of a natural extract from Mangifera indica L (Vimang), its main ingredient mangiferin and epigallocatechin gallate (EGCG) on energy metabolism, energy state and malondialdehyde (MDA) production in a red blood cell system. Analysis of MDA, high energy phosphates and ascorbate was carried out by high performance liquid chromatography (HPLC). Under the experimental conditions, concentrations of MDA and ATP catabolites were affected in a dose-dependent way by H2O2. Incubation with Vimang (0.1, 1, 10, 50 and 100 microg/mL), mangiferin (1, 10, 100 microg/mL) and EGCG (0.01, 0.1, 1, 10 microM) significantly enhances erythrocyte resistance to H2O2-induced reactive oxygen species production. In particular, we demonstrate the protective activity of these compounds on ATP, GTP and total nucleotides (NT) depletion after H2O2-induced damage and a reduction of NAD and ADP, which both increase because of the energy consumption following H2O2 addition. Energy charge potential, decreased in H2O2-treated erythrocytes, was also restored in a dose-dependent way by these substances. Their protective effects might be related to the strong free radical scavenging ability described for polyphenols.

Topics: Ascorbic Acid; Catechin; Chromatography, High Pressure Liquid; Erythrocytes; Hemolysis; Humans; Hydrogen Peroxide; Lipid Peroxidation; Malondialdehyde; Mangifera; Plant Extracts; Xanthones

One of the methods of increasing the quality of fatty products is including in their composition antioxidants, which provide the time of their conservation and improvement of their biological value. From this point of view calls concern a new drug epigallocatechina gallate. On the basis of these studies the prospects of usage were shown epigallocatechina gallate in fatty products and the paths of further applying in creation of food products.

Topics: alpha-Tocopherol; Antioxidants; Ascorbic Acid; Butylated Hydroxyanisole; Catechin; Cinnamates; Depsides; Dietary Fats; Emulsions; Food Preservation; Plants; Rosmarinic Acid; Soybean Oil

Current treatment of osteosarcoma is associated with poor prognosis, especially due to the increased risk of developing other cancers with chemotherapy. Therefore, new, safe and effective treatment strategies are needed. We investigated the effect of a unique mixture of nutrients containing lysine, proline, arginine, ascorbic acid, and epigallocatechin gallate (EGCG) on human osteosarcoma cell lines U-2OS, MNNG-HOS, and Ewing's sarcoma SK-ES-1 by measuring: cell proliferation, expression of matrix metalloproteinase-2 (MMP-2), MMP-9, and invasive and angiogenesis potential. Cell proliferation was evaluated by MTT assay, matrix metalloproteinases (MMP) expression by gelatinase zymography, VEGF expression by ELISA, and invasion through Matrigel. Cells were also treated with phorbol 12-myristate 13-acetate (PMA) to study enhanced MMP and VEGF expression. The invasion of osteosarcoma U-2OS and MNNG-HOS cells through Matrigel was significantly reduced in a dose-dependent fashion, with 100% inhibition of invasion of U-2OS cells at 100 microg/ml, and MNNG cells at 50 microg/ml concentration of the synergistically acting nutrient mixture. Ewing's sarcoma SK-ES-1 cells were not invasive. Nutrient synergy (NS) exhibited a dose response antiproliferative effect on osteosarcoma U-2OS cells, reaching 67% at 1000 microg/ml of NS; no significant suppression of cell proliferation was seen with MNNG or Ewing's sarcoma cells. Zymography showed dose-dependent inhibition of MMP secretion by all three cell lines in the presence of NS. VEGF secretion by U-2OS cells was completely blocked at 500 microg/ml of NS. Our results suggest NS is an excellent candidate for therapeutic use in the treatment of osteosarcoma, by inhibiting cancer cell invasion, and secretion of MMPs and VEGF, all critical parameters for cancer control and prevention.

Topics: Arginine; Ascorbic Acid; Bone Neoplasms; Catechin; Cell Proliferation; Dose-Response Relationship, Drug; Humans; Lysine; Matrix Metalloproteinases; Neovascularization, Pathologic; Nutritional Physiological Phenomena; Osteosarcoma; Proline; Sarcoma, Ewing; Tetradecanoylphorbol Acetate; Tumor Cells, Cultured; Vascular Endothelial Growth Factor A

The ability of various dietary compounds to modulate the activity of the transcription factor nuclear factor kappaB (NF-kappaB) was examined using a cell-based reporter system. NF-kappaB is central to the response of cells to stress and has been linked to cancer. HCT 116 (human colon carcinoma) and HepG2 (human liver carcinoma) cell lines were stably transfected with a NF-kappaB luciferase reporter vector. The reporter cell lines were preincubated with different concentrations (0-50 microM) of ascorbic acid, epigallocatechin gallate, genistein, quercetin, naringenin, and resveratrol for varying periods of times (1-12 hours), after which the NF-kappaB inducer tumor necrosis factor-alpha (TNF-alpha) was added (4-8 ng/mL) for 4 hours. Compound alone, without TNF-alpha, did not alter luciferase activity. Levels of TNF-alpha-induced luciferase (NF-kappaB) activity varied depending on compound type and concentration, whereas preincubation time and cell type contributed less. Significant changes in luciferase (NF-kappaB) activity were detected for some of the compounds at more physiological concentrations (1-10 microM). Our data suggest that dietary modulation of NF-kappaB activity involves distinct mechanisms, depending on compound type and concentration. More generally, this approach can be utilized for analyzing dietary compounds for effects on specific cellular factors over a range of concentrations and incubation times, in combination, and in different cell types.

Topics: Antioxidants; Ascorbic Acid; Catechin; Dose-Response Relationship, Drug; Flavonoids; HCT116 Cells; Humans; Luciferases; NF-kappa B; Time Factors; Tumor Cells, Cultured; Tumor Necrosis Factor-alpha

The combination of indole-3-acetic acid (IAA) and horseradish peroxidase (HRP) has recently been proposed as a novel cancer therapy. However, the mechanism underlying the cytotoxic effect involved is substantially unknown. Here, we show that IAA/HRP treatment induces apoptosis in G361 human melanoma cells, whereas IAA or HRP alone have no effect. It is known that IAA produces free radicals when oxidized by HRP. Because oxidative stress could induce apoptosis, we measured the production of free radicals at varying concentrations of IAA and HRP. Our results show that IAA/HRP produces free radicals in a dose-dependent manner, which are suppressed by ascorbic acid or (-)-epigallocatechin gallate (EGCG). Furthermore, antioxidants prevent IAA/HRP-induced apoptosis, indicating that the IAA/HRP-produced free radicals play an important role in the apoptotic process. In addition, IAA/HRP was observed to activate p38 mitogen-activated protein (MAP) kinase and c-Jun N-terminal kinase (JNK), which are almost completely blocked by antioxidants. We further investigated the IAA/HRP-mediated apoptotic pathways, and found that IAA/HRP activates caspase-8 and caspase-9, leading to caspase-3 activation and poly(ADP-ribose) polymerase (PARP) cleavage. These events were also blocked by antioxidants, such as ascorbic acid or EGCG. Thus, we propose that IAA/HRP-induced free radicals lead to the apoptosis of human melanoma cells via both death receptor-mediated and mitochondrial apoptotic pathways.

Topics: Antioxidants; Apoptosis; Ascorbic Acid; Caspases; Catechin; Cell Line, Tumor; Dose-Response Relationship, Drug; Drug Screening Assays, Antitumor; Drug Synergism; Drug Therapy, Combination; Free Radicals; Horseradish Peroxidase; Humans; Indoleacetic Acids; JNK Mitogen-Activated Protein Kinases; Melanoma; Mitochondria; Mitogen-Activated Protein Kinases; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Poly (ADP-Ribose) Polymerase-1; Poly(ADP-ribose) Polymerases; Prodrugs; Proteins

Once prostate cancer has metastasized, current treatment methods are generally ineffective. Due to the reported anti-tumor properties of specific nutrients, we investigated the effect of a unique formulation (NS) of lysine, proline, arginine, ascorbic acid, and epigallocatechin gallate on human prostate cancer cell lines: PC-3, DU145 (androgen insensitive) and LNCaP (androgen sensitive), by measuring cell proliferation, MMP expression, and invasion potential. Cell lines DU145, PC-3, and LNCaP were treated at near confluence with NS at various concentrations. Cell proliferation was measured by MTT assay after 24 hours, MMP expression was measured by gelatinase zymography in condition media, and invasion activity was measured by Matrigel. The nutrient mixture did not significantly inhibit PC-3 cell proliferation at 50 microg/ml, but showed significant antiproliferative effect at 500 ug/ml. When treated with NS, proliferation of LNCaP cells was inhibited by 80% of control at 100 microg/ml. NS showed dose-dependent inhibition of DU145 cell proliferation with 47% reduction at 1000 microg/ml. NS showed a dose-dependent inhibition of both MMP-2 and MMP-9 expression by PC-3 cells and MMP-9 expression by PMA-treated (200 ng/ml) DU145 cells. Neither MMP-2 nor MMP-9 gelatinolytic activity was detected in LNCaP cell culture. Invasion of DU145 and LNCaP cells through Matrigel was completely inhibited at 500 microg/ml and PC-3 at 1000 microg/ml. Inhibition of MMP expression and invasion suggests the mixture of nutrients studied is a potent, natural anticancer agent for the treatment of prostate cancer.

Topics: Antioxidants; Arginine; Ascorbic Acid; Catechin; Cell Line, Tumor; Cell Proliferation; Chemistry, Pharmaceutical; Collagen; Dose-Response Relationship, Drug; Drug Combinations; Drug Synergism; Formazans; Humans; Laminin; Lysine; Male; Matrix Metalloproteinases; Neoplasm Invasiveness; Proline; Prostatic Neoplasms; Proteoglycans; Tetrazolium Salts

DNA damage generated by oxygen-derived free radicals is related to mutagenesis, carcinogenesis and aging. In the last several years, hundreds of publications have confirmed that melatonin is a potent endogenous free radical scavenger. In the present in vitro study, we have examined the efficacy of three polyphenolic antioxidants, i.e. xanthurenic acid, resveratrol (3,4',5-trihydroxy-trans-stilbene) and (-)-epigallocatechin-3-gallate (EGCG) and two classical non-polyphenolic antioxidants, i.e. vitamin C (ascorbic acid) and alpha-lipoic acid (LA, 1,2-dithiolane-3-pentanoic acid) in inhibiting *OH-induced oxidative DNA damage. We compared the efficacy of these five antioxidants with the effectiveness of melatonin (N-acetyl-5-methoxytryptamine) and we also investigated the possible synergistic effects of melatonin with the other five molecules. Using high performance liquid chromatography (HPLC), the formation of 8-hydroxy-2-deoxyguanosine (8-OH-dG) in purified calf thymus DNA treated with the Fenton reagents, chromium(III) (as CrCl3) plus hydrogen peroxide (H2O2) (Cr(III)/H2O2), was measured in the presence or absence of the antioxidants alone or in combination with melatonin. 8-OH-dG is considered a biomarker of oxidative DNA damage. Among the antioxidants tested, melatonin was the most effective of these with an IC50 = 3.6 +/- 0.1 micro m. For the other antioxidants the IC50 values were as follows: xanthurenic acid (IC50 = 7.9 +/- 0.3), resveratrol (IC50 = 10.9 +/- 0.3), EGCG (IC50 = 5.7 +/- 0.3), vitamin C (IC50 = 16.9 +/- 0.5) and LA (IC50 = 38.8 +/- 0.7). These values differ from that of melatonin with a P < 0.01. Melatonin (1 micro M) reversed the pro-oxidant effect of resveratrol (0.5 micro M) and vitamin C (0.5 micro M), had an antagonistic effect when used in combination with EGCG (1 micro M) and it exhibited synergism in combination with vitamin C (0.5 micro M) and with LA (5 micro M).

Topics: 8-Hydroxy-2'-Deoxyguanosine; Antioxidants; Ascorbic Acid; Catechin; Chromium; Deoxyguanosine; DNA Damage; Dose-Response Relationship, Drug; Drug Synergism; Hydrogen Peroxide; Inhibitory Concentration 50; Iron; Melatonin; Oxidative Stress; Resveratrol; Stilbenes; Thioctic Acid; Xanthurenates

A mixture of (-)-epigallocatechin-3-gallate (EGCG) and ascorbic acid exhibited 73.2% inhibition of SMMC-7721 cell proliferation in a soft agar colony formation assay, which was much higher than EGCG (40.4%) or ascorbic acid (12.4%) alone. In the cell migration assay, the mixture also significantly suppressed the migration of SMMC-7721 cells by 65.9% while EGCG and/or ascorbic acid did by 28.9% and 18.7%, respectively. Ascorbic acid was able to enhance the antioxidant activity of EGCG by decreasing the intracellular oxidative stress according to fluorographic analysis of oxidative stress. In conclusion, the combination of EGCG and ascorbic acid can strongly suppress the proliferation and metastasis of liver cancer cells, possibly with a mechanism associated with the scavenging of reactive oxygen species. All these events add to our knowledge of liver cancer chemotherapy.

Topics: Antioxidants; Ascorbic Acid; Catechin; Cell Division; Cell Movement; Colony Count, Microbial; Culture Media; Drug Therapy, Combination; Humans; Liver Neoplasms; Reference Values; Sensitivity and Specificity; Tumor Cells, Cultured

Phenolic compounds have recently attracted special attention due to their beneficial health effects; their intestinal absorption and bioavailability need, therefore, to be investigated and Caco-2 cell culture model appeared as a promising tool. We have shown herein that the addition of a grape seed extract (GSE) to Dulbecco's modified Eagle's medium (DMEM) used for Caco-2 cell culture leads to a substantial loss of catechin, epicatechin and B2 and B3 dimers from GSE in the medium after 24 h and to a production of hydrogen peroxide (H2O2). When 1420 microM ascorbic acid is added to the DMEM, such H2O2 production was prevented. This hydrogen peroxide generation substantially involves inorganic salts from the DMEM. We recommend that ascorbic acid be added to circumvent such a risk.

Topics: Antioxidants; Ascorbic Acid; Caco-2 Cells; Catechin; Culture Media; Humans; Hydrogen Peroxide; Vitis

The purpose of this study was to investigate the effects of single/double or repeated intake of a normal amount of tea catechin on plasma catechin concentrations and antioxidant activity in young women.. First, after an overnight fast, five healthy subjects were given water or single/double dose(s) of tea polyphenol extract (164 mg tea catechins containing 61% epigallocatechin gallate in 190 ml water). Blood samples were taken before and 30, 60 and 180 min after the ingestion. Second, 16 healthy subjects ingested the tea polyphenol extract three times a day at mealtimes for 7 days followed by withdrawal of tea polyphenol extract for 7 days. Blood samples were taken before and after ingestion, and 7 days after the withdrawal of tea catechin. Subjects were prohibited from drinking any beverages containing polyphenols or antioxidant supplements during the study period. Catechin and other antioxidant concentrations in the plasma were measured, and changes in antioxidant activity were evaluated by ferric reducing ability of plasma assay.. Single/double ingestion of tea polyphenol extract did not cause an increase in the antioxidant activity. There was no also change in antioxidant activity after the ingestion of tea polyphenol extract for 7 days. Plasma-free epigallocatechin gallate concentration remained at the pre-study level; however, the plasma FRAP value decreased significantly at 7 days after the withdrawal of tea polyphenol extract. Decreases in endogenous antioxidants in the plasma, including vitamin C and bilirubin, were also observed 7 days after withdrawal of tea polyphenol.. The results suggest that continuous daily intake of tea catechins affects the concentrations of endogenous antioxidants in the plasma and has the potential to maintain total antioxidant activity.

Topics: Adult; alpha-Tocopherol; Antioxidants; Ascorbic Acid; Bilirubin; Catechin; Cholesterol; Dose-Response Relationship, Drug; Female; Flavonoids; Humans; Kinetics; Phenols; Plant Extracts; Polymers; Tea; Uric Acid

The protective effect of fruits and vegetables against cancer is well established. It is believed that this effect is mediated by antioxidants and decreased oxidative damage to DNA. However, the identity of the antioxidant(s) responsible is not clear. Moreover, a potentially damaging pro-oxidant effect of some antioxidants has been reported. In this study the ex vivo effects of several dietary antioxidants, including quercetin, various catechins, ascorbic acid and alpha-tocopherol, were investigated, at concentrations up to 200 microM, using the single cell gel electrophoresis (comet) assay for DNA damage. Lymphocytes from three healthy subjects were pre-incubated with these antioxidants, and the comet assay was performed on treated, untreated, challenged and unchallenged cells in parallel, oxidant challenge being induced by 5 min exposure to hydrogen peroxide (final concentrations H2O2: 30, 45, or 60 microM). Results using this ex vivo cellular assay showed protection by some antioxidants (quercetin, caffeic acid), no effect by some (catechin, epicatechin, catechin gallate, epicatechin gallate) and an apparently damaging effect by others (epigallocatechin, epigallocatechin gallate). Damage may have been caused by production of H2O2 from these polyphenolics. Neither ascorbic acid nor alpha-tocopherol protected or damaged DNA. Further study of the role of quercetin and caffeic acid in DNA protection is needed.

Topics: alpha-Tocopherol; Antioxidants; Ascorbic Acid; Caffeic Acids; Catechin; Cells, Cultured; Comet Assay; DNA; DNA Damage; Dose-Response Relationship, Drug; Female; Free Radical Scavengers; Humans; Hydrogen Peroxide; Lymphocytes; Male; Oxidative Stress; Quercetin; Time Factors

Both ascorbic acid and epigallocatechin gallate, which is one of the key polyphenols contained in green tea leaves, have been considered excellent antioxidants. The present study compared the efficacy as antioxidants between the two agents on an equimolar basis. Cells of two lymphoid lines were used as test material to determine the reduction of chromosome damage induced by the radiomimetic antibiotic bleomycin. Without bleomycin, both agents, at concentrations of 10(-7), 10(-6), 10(-5), and 10(-4) M, showed chromosome damage similar to the untreated controls. With bleomycin, the weakest concentration of both showed no protective effect. At concentrations of 10(-6) and 10(-5) M, especially the latter, a significant reduction in frequencies of chromatid breaks was recorded. However, at the highest concentration, 10(-4) M, the chromatid break frequencies rose to the same level as that of cells treated with bleomycin alone, suggesting that both behaved like pro-oxidants.

Topics: Antioxidants; Ascorbic Acid; Bleomycin; Catechin; Cell Line; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Humans; Lymphocytes; Mutagens

Cells lacking mismatch repair (MMR) exhibit elevated levels of spontaneous mutagenesis. Evidence exists that MMR is involved in repair of some DNA lesions besides mismatches. If some oxidative DNA lesions are substrates for MMR, then the excess mutagenesis in MMR(-) cells might be blocked by dietary antioxidants. Effects of the dietary antioxidants ascorbate, alpha-tocopherol, (-)-epigallocatechin gallate (EGCG) and lycopene on spontaneous mutagenesis were studied using mismatch repair-deficient (hMLH1(-)) human colon carcinoma HCT116 cells and HCT116/ch3 cells, in which normal human chromosome 3 has been added to restore mismatch repair. HCT116 cells have a 22-fold higher spontaneous mutation rate compared with HCT116/ch3 cells. HCT116 cells cultured in 1% fetal bovine serum (FBS) have twice the spontaneous mutation rate of those cultured in 10% FBS, most likely due to reduction in serum antioxidants in the low serum medium. As expected, alpha-tocopherol (50 microM) and ascorbate (284 microM) reduced spontaneous mutagenesis in HCT116 cells growing in 1% serum more dramatically than in cells cultured in 10% serum. The strongest antimutagenic compound was lycopene (5 microM), which reduced spontaneous mutagenesis equally (about 70%) in HCT116 cells growing in 10 and 1% FBS and in HCT116/ch3 cells. Since lycopene was equally antimutagenic in cells growing in low and high serum, it may have another antimutagenic mechanism in addition to its antioxidant effect. Surprisingly, EGCG (10 microM) was toxic to cells growing in low serum. It also reduced spontaneous mutagenesis equally (nearly 40%) in HCT116 and HCT116/ch3 cells. The large proportion of spontaneous mutagenesis that can be blocked by antioxidants in mismatch repair-deficient cells support the hypothesis that a major cause of their excess mutagenesis is endogenous oxidants. Blocking spontaneous mutagenesis, perhaps with a cocktail of antioxidants, should reduce the risk of cancer in people with a genetic defect in mismatch repair as well as other individuals.

Topics: Antioxidants; Ascorbic Acid; Base Pair Mismatch; Blood Proteins; Carotenoids; Catechin; Cell Line; Cell Survival; Chromosomes, Human, Pair 3; Clone Cells; Dietary Supplements; DNA Repair; Humans; Lycopene; Mutagenesis; Mutagenicity Tests; Vitamin E

Topics: Antioxidants; Ascorbic Acid; Catechin; Chromans; DNA Damage; DNA, Bacterial; Flavonoids; Gamma Rays; Glutathione; Hydroxyl Radical; Models, Biological; Oxidants; Oxidation-Reduction

Exposure of normal human epidermal keratinocytes (NHEK) to UVB radiation induces intracellular release of hydrogen peroxide (oxidative stress) and phosphorylation of mitogen-activated protein kinase cell signaling pathways. Here, we demonstrate that pretreatment of NHEK with (-)-epigallocatechin-3-gallate (EGCG), an antioxidant from green tea, inhibits UVB-induced hydrogen peroxide (H(2)O(2)) production and H(2)O(2)-mediated phosphorylation of MAPK signaling pathways. We found that treatment of EGCG (20 microg/ml of media) to NHEK before UVB (30 mJ/cm(2)) exposure inhibited UVB-induced H(2)O(2) production (66-80%) concomitant with the inhibition of UVB-induced phosphorylation of ERK1/2 (57-80%), JNK (53-83%), and p38 (50-77%) proteins. To demonstrate whether UVB-induced phosphorylation of MAPK occurs via UVB-induced H(2)O(2) (oxidative stress) production, NHEK were treated with the oxidant H(2)O(2). Treatment of H(2)O(2) to NHEK resulted in phosphorylation of ERK1/2, JNK, and p38. Using the same in vitro system, when these cells were pretreated with EGCG or with the known antioxidant ascorbic acid (as positive control), H(2)O(2)-induced phosphorylation of ERK1/2, JNK, and p38 was found to be significantly inhibited. These findings demonstrate that EGCG has the potential to inhibit UVB-induced oxidative stress-mediated phosphorylation of MAPK signaling pathways, suggesting that EGCG could be useful in attenuation of oxidative stress-mediated and MAPK-caused skin disorders in humans.

Topics: Antioxidants; Ascorbic Acid; Blotting, Western; Catechin; Epidermal Cells; Epidermis; Humans; Hydrogen Peroxide; JNK Mitogen-Activated Protein Kinases; Keratinocytes; MAP Kinase Kinase 4; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 3; Mitogen-Activated Protein Kinase Kinases; Mitogen-Activated Protein Kinases; Oxidants; Oxidative Stress; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Ultraviolet Rays

Blood plasma was incubated with 50 mM AAPH [2, 2'-azobis-(2-amidinopropane) hydrochloride] in the absence or presence of catechins (5-100 microM). Lipid oxidation was evaluated by measuring the formation of 2-thiobarbituric acid reactive substances (TBARS). The concentration of alpha-tocopherol (AT), beta-carotene (BC), ascorbic acid (AA), and catechins was determined by reverse phase high performance liquid chromatography (HPLC) with electrochemical detection. All the assayed catechins inhibited plasma TBARS formation. Based on the calculated IC50, the order of effectiveness was: epicatechin gallate (ECG) > epigallocatechin gallate (EGCG) > epigallocatechin (EGC) > epicatechin (EC) > catechin (C). Catechins protected plasma AT and BC from AAPH-mediated oxidation. The order of effectiveness for AT protection was ECG > EGCG > EC = C > EGC; and for BC protection, the order was EGCG > ECG > EGC > > EC > C. The addition of catechins modified the kinetics of TBARS formation and AT depletion, but the rate of AA depletion was not affected. Catechin oxidation did not start until the complete depletion of AA, and it preceded AT depletion. These results indicate that catechins are effective antioxidants in human blood plasma, delaying the lipid oxidation and depletion of endogenous lipid-soluble antioxidants (AT and BC).

Topics: Ascorbic Acid; beta Carotene; Catechin; Chromatography, High Pressure Liquid; Dose-Response Relationship, Drug; Flavonoids; Humans; Kinetics; Lipid Peroxidation; Thiobarbituric Acid Reactive Substances; Vitamin E

Gamma ray-induced scission of puC18 plasmid DNA prepared from E. coli was examined in the presence of a green tea percolate and its main constituents, L-ascorbic acid (used as the sodium salt) and (-)-epigallocatechin gallate. Each of these showed a protecting effect against DNA scission. The relationship between the protecting effect against DNA scission and the scavenging effect of the hydroxyl radical was examined, and is discussed from the viewpoint of interaction with DNA.

Topics: Antimutagenic Agents; Ascorbic Acid; Catechin; DNA Damage; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Escherichia coli; Free Radical Scavengers; Gamma Rays; Hydroxyl Radical; Plant Extracts; Plasmids; Stereoisomerism; Tea